Genetic elements, proteins, and associated methods including application of additional genetic information to gram (+) thermoacidophiles

ABSTRACT

Isolated and/or purified polypeptides and nucleic acid sequences encoding polypeptides from  Alicyclobacillus acidocaldarius  are provided. Further provided are methods for modulating or altering recombination inside or outside of a cell using isolated and/or purified polypeptides and/or nucleic acid sequences from  Alicyclobacillus acidocaldarius.

GOVERNMENT RIGHTS

The United States Government has certain rights in this inventionpursuant to Contract No. DE-AC07-99ID13727 and Contract No.DE-AC07-05ID14517 between the United States Department of Energy andBattelle Energy Alliance, LLC.

STATEMENT ACCORDING TO 37 C.F.R. §1.52(e)(5)-SEQUENCE LISTING SUBMITTEDON COMPACT DISC

Pursuant to 37 C.F.R. §1.52(e)(1)(ii), a compact disc containing anelectronic version of the Sequence Listing has been submittedconcomitant with this application, the contents of which are herebyincorporated by reference. A second compact disc is submitted and is anidentical copy of the first compact disc. The discs are labeled “copy 1”and “copy 2,” respectively, and each disc contains one file entitled“Seq list BA 315.txt” which is 964 KB and created on January XX, 2010.

TECHNICAL FIELD

The present invention relates generally to biotechnology. Morespecifically, embodiments of the present invention relate to isolatedand/or purified polypeptides and nucleic acid sequences encodingpolypeptides from Alicyclobacillus acidocaldarius and methods for theiruse.

BACKGROUND

Thermophilic and/or acidophilic bacteria have great potential forproduction of useful chemicals in industrial processes. However, mostsystems for promoting genetic recombination for the purposes ofintroducing nucleic acids of interest are not well suited for suchthermophilic and/or acidophilic bacteria.

BRIEF SUMMARY OF THE INVENTION

Embodiments of the invention relate to purified and/or isolatednucleotide sequences of the genome of Alicyclobacillus acidocaldarius,or a homologue or fragment thereof. In one embodiment of the invention,the nucleotide sequence is selected from at least one of SEQ ID Nos. 2,19, 36, 53, 70, 87, 104, 121, 138, 155, 172, 189, 206, 223, 240, 257,274, 291, 308, 325, 359, 376, 410, 427, 444, 512, 529, 546, 597, 614,648, 665, 682, 699, 716, 733, 750, 767, 784, 818, 835, 852, 886, 903,920, 937, 954, 971, 988, 1005, 1039, 1056, 1073, 1090, 1107, 1124, 1141,1158, 1175, 1192, 1209, 1226, 1243, and 1260, 1277, 1294, 1311, 1328,1345, 1362, 1379, 1396, 1413, 1430, 1447, 1464, 1481, 1498, 1515, 1532,1549, 1566, 1583, 1600, 1634, 1651, 1685, 1702, 1719, 1787, 1804, 1821,1872, 1889, 1923, 1940, 1957, 1974, 1991, 2008, 2025, 2042, 2059, 2093,2110, 2127, 2161, 2178, 2195, 2212, 2229, 2246, 2263, 2280, 2314, 2331,2348, 2365, 2382, 2399, 2416, 2433, 2450, 2467, 2484, 2501, 2518, 2535,2552, 2569, 2586, 2603, 2620, 2637, 2654, 2671, 2688, 2705, 2722, 2739,2773, 2790, 2824, 2841, 2858, 2926, 2943, 2960, 3011, 3028, 3062, 3079,3096, 3113, 3130, 3147, 3164, 3181, 3198, 3232, 3249, 3266, and 3283 ora homologue or fragment thereof. In another embodiment of the invention,the homologue is selected from the group consisting of a nucleotidesequence having at least 80% sequence identity to at least one of SEQ IDNos. 2, 19, 36, 53, 70, 87, 104, 121, 138, 155, 172, 189, 206, 223, 240,257, 274, 291, 308, 325, 359, 376, 410, 427, 444, 512, 529, 546, 597,614, 648, 665, 682, 699, 716, 733, 750, 767, 784, 818, 835, 852, 886,903, 920, 937, 954, 971, 988, 1005, 1039, 1056, 1073, 1090, 1107, 1124,1141, 1158, 1175, 1192, 1209, 1226, 1243, 1260, 1277, 1294, 1311, 1328,1345, 1362, 1379, 1396, 1413, 1430, 1447, 1464, 1481, 1498, 1515, 1532,1549, 1566, 1583, 1600, 1634, 1651, 1685, 1702, 1719, 1787, 1804, 1821,1872, 1889, 1923, 1940, 1957, 1974, 1991, 2008, 2025, 2042, 2059, 2093,2110, 2127, 2161, 2178, 2195, 2212, 2229, 2246, 2263, 2280, 2314, 2331,2348, 2365, 2382, 2399, 2416, 2433, 2450, 2467, 2484, 2501, 2518, 2535,2552, 2569, 2586, 2603, 2620, 2637, 2654, 2671, 2688, 2705, 2722, 2739,2773, 2790, 2824, 2841, 2858, 2926, 2943, 2960, 3011, 3028, 3062, 3079,3096, 3113, 3130, 3147, 3164, 3181, 3198, 3232, 3249, 3266, and 3283.

Embodiments of the invention may further relate to an isolated and/orpurified nucleic acid sequence comprising a nucleic acid sequenceencoding a polypeptide selected from the group consisting of apolypeptide having at least 90% sequence identity to at least one of SEQID Nos. 1, 18, 35, 52, 69, 86, 103, 120, 137, 154, 171, 188, 205, 222,239, 256, 273, 290, 307, 324, 358, 375, 409, 426, 443, 511, 528, 545,596, 613, 647, 664, 681, 698, 715, 732, 749, 766, 783, 817, 834, 851,885, 902, 819, 936, 953, 970, 987, 1004, 1038, 1055, 1072, 1089, 1106,1123, 1140, 1157, 1174, 1191, 1208, 1225, 1242, 1259, 1276, 1293, 1310,1327, 1344, 1361, 1378, 1395, 1412, 1429, 1446, 1463, 1480, 1497, 1514,1531, 1548, 1565, 1582, 1599, 1633, 1650, 1684, 1701, 1718, 1786, 1803,1820, 1871, 1888, 1922, 1939, 1956, 1973, 1990, 2007, 2024, 2041, 2058,2092, 2109, 2126, 2160, 2177, 2094, 2211, 2228, 2245, 2262, 2279, 2313,2330, 2347, 2364, 2381, 2398, 2415, 2432, 2449, 2466, 2483, 2500, 2517,2534, 2551, 2568, 2585, 2602, 2619, 2636, 2653, 2670, 2687, 2704, 2721,2738, 2772, 2789, 2823, 2840, 2857, 2925, 2942, 2959, 3010, 3027, 3061,3078, 3095, 3112, 3129, 3146, 3163, 3180, 3197, 3231, 3248, 3265, and3282.

Embodiments of the invention also relate to isolated and/or purifiedpolypeptides coded for by a nucleotide sequence comprising a nucleotidesequence of the genome of Alicyclobacillus acidocaldarius, or ahomologue or fragment thereof. In one embodiment, the nucleotidesequence comprises a nucleotide sequence selected from the groupconsisting of a nucleotide sequence having at least 80% sequenceidentity to at least one of SEQ ID Nos. 2, 19, 36, 53, 70, 87, 104, 121,138, 155, 172, 189, 206, 223, 240, 257, 274, 291, 308, 325, 359, 376,410, 427, 444, 512, 529, 546, 597, 614, 648, 665, 682, 699, 716, 733,750, 767, 784, 818, 835, 852, 886, 903, 920, 937, 954, 971, 988, 1005,1039, 1056, 1073, 1090, 1107, 1124, 1141, 1158, 1175, 1192, 1209, 1226,1243, 1260, 1277, 1294, 1311, 1328, 1345, 1362, 1379, 1396, 1413, 1430,1447, 1464, 1481, 1498, 1515, 1532, 1549, 1566, 1583, 1600, 1634, 1651,1685, 1702, 1719, 1787, 1804, 1821, 1872, 1889, 1923, 1940, 1957, 1974,1991, 2008, 2025, 2042, 2059, 2093, 2110, 2127, 2161, 2178, 2195, 2212,2229, 2246, 2263, 2280, 2314, 2331, 2348, 2365, 2382, 2399, 2416, 2433,2450, 2467, 2484, 2501, 2518, 2535, 2552, 2569, 2586, 2603, 2620, 2637,2654, 2671, 2688, 2705, 2722, 2739, 2773, 2790, 2824, 2841, 2858, 2926,2943, 2960, 3011, 3028, 3062, 3079, 3096, 3113, 3130, 3147, 3164, 3181,3198, 3232, 3249, 3266, and 3283.

In another embodiment of the invention, the nucleotide sequencecomprises a nucleotide sequence selected from at least one of SEQ IDNos. 2, 19, 36, 53, 70, 87, 104, 121, 138, 155, 172, 189, 206, 223, 240,257, 274, 291, 308, 325, 359, 376, 410, 427, 444, 512, 529, 546, 597,614, 648, 665, 682, 699, 716, 733, 750, 767, 784, 818, 835, 852, 886,903, 920, 937, 954, 971, 988, 1005, 1039, 1056, 1073, 1090, 1107, 1124,1141, 1158, 1175, 1192, 1209, 1226, 1243, and 1260, 1277, 1294, 1311,1328, 1345, 1362, 1379, 1396, 1413, 1430, 1447, 1464, 1481, 1498, 1515,1532, 1549, 1566, 1583, 1600, 1634, 1651, 1685, 1702, 1719, 1787, 1804,1821, 1872, 1889, 1923, 1940, 1957, 1974, 1991, 2008, 2025, 2042, 2059,2093, 2110, 2127, 2161, 2178, 2195, 2212, 2229, 2246, 2263, 2280, 2314,2331, 2348, 2365, 2382, 2399, 2416, 2433, 2450, 2467, 2484, 2501, 2518,535, 2552, 2569, 2586, 2603, 2620, 2637, 2654, 2671, 2688, 2705, 2722,2739, 2773, 2790, 2824, 2841, 2858, 2926, 2943, 2960, 3011, 3028, 3062,3079, 3096, 3113, 3130, 3147, 3164, 3181, 3198, 3232, 3249, 3266, and3283 or a homologue or fragment thereof In still another embodiment, thepolypeptide comprises an amino acid sequence of SEQ ID Nos. 1, 18, 35,52, 69, 86, 103, 120, 137, 154, 171, 188, 205, 222, 239, 256, 273, 290,307, 324, 358, 375, 409, 426, 443, 511, 528, 545, 596, 613, 647, 664,681, 698, 715, 732, 749, 766, 783, 817, 834, 851, 885, 902, 819, 936,953, 970, 987, 1004, 1038, 1055, 1072, 1089, 1106, 1123, 1140, 1157,1174, 1191, 1208, 1225, 1242, 1259, 1276, 1293, 1310, 1327, 1344, 1361,1378, 1395, 1412, 1429, 1446, 1463, 1480, 1497, 1514, 1531, 1548, 1565,1582, 1599, 1633, 1650, 1684, 1701, 1718, 1786, 1803, 1820, 1871, 1888,1922, 1939, 1956, 1973, 1990, 2007, 2024, 2041, 2058, 2092, 2109, 2126,2160, 2177, 2094, 2211, 2228, 2245, 2262, 2279, 2313, 2330, 2347, 2364,2381, 2398, 2415, 2432, 2449, 2466, 2483, 2500, 2517, 2534, 2551, 2568,2585, 2602, 2619, 2636, 2653, 2670, 2687, 2704, 2721, 2738, 2772, 2789,2823, 2840, 2857, 2925, 2942, 2959, 3010, 3027, 3061, 3078, 3095, 3112,3129, 3146, 3163, 3180, 3197, 3231, 3248, 3265, and 3282. In yet anotherembodiment, the polypeptide comprises an amino acid sequence selectedfrom the group consisting of a polypeptide having at least 90% sequenceidentity to at least one of SEQ ID Nos. 1, 18, 35, 52, 69, 86, 103, 120,137, 154, 171, 188, 205, 222, 239, 256, 273, 290, 307, 324, 358, 375,409, 426, 443, 511, 528, 545, 596, 613, 647, 664, 681, 698, 715, 732,749, 766, 783, 817, 834, 851, 885, 902, 819, 936, 953, 970, 987, 1004,1038, 1055, 1072, 1089, 1106, 1123, 1140, 1157, 1174, 1191, 1208, 1225,1242, 1259, 1276, 1293, 1310, 1327, 1344, 1361, 1378, 1395, 1412, 1429,1446, 1463, 1480, 1497, 1514, 1531, 1548, 1565, 1582, 1599, 1633, 1650,1684, 1701, 1718, 1786, 1803, 1820, 1871, 1888, 1922, 1939, 1956, 1973,1990, 2007, 2024, 2041, 2058, 2092, 2109, 2126, 2160, 2177, 2094, 2211,2228, 2245, 2262, 2279, 2313, 2330, 2347, 2364, 2381, 2398, 2415, 2432,2449, 2466, 2483, 2500, 2517, 2534, 2551, 2568, 2585, 2602, 2619, 2636,2653, 2670, 2687, 2704, 2721, 2738, 2772, 2789, 2823, 2840, 2857, 2925,2942, 2959, 3010, 3027, 3061, 3078, 3095, 3112, 3129, 3146, 3163, 3180,3197, 3231, 3248, 3265, and 3282.

In embodiments of the invention, the polypeptides may be acidophilicand/or thermophilic. In further embodiments, the polypeptides may beglycosylated, pegylated, and/or otherwise post-translationally modified.

Embodiments of methods include methods of altering genetic recombinationinside or outside of a cell, the methods comprising providing arecombinant, purified, and/or isolated nucleotide sequence comprising anucleotide sequence selected from the group consisting of a nucleotidesequences having at least 90% sequence identity to at least one of thesequences of SEQ ID Nos. 2, 19, 36, 53, 70, 87, 104, 121, 138, 155, 172,189, 206, 223, 240, 257, 274, 291, 308, 325, 359, 376, 410, 427, 444,512, 529, 546, 597, 614, 648, 665, 682, 699, 716, 733, 750, 767, 784,818, 835, 852, 886, 903, 920, 937, 954, 971, 988, 1005, 1039, 1056,1073, 1090, 1107, 1124, 1141, 1158, 1175, 1192, 1209, 1226, 1243, 1260,1277, 1294, 1311, 1328, 1345, 1362, 1379, 1396, 1413, 1430, 1447, 1464,1481, 1498, 1515, 1532, 1549, 1566, 1583, 1600, 1634, 1651, 1685, 1702,1719, 1787, 1804, 1821, 1872, 1889, 1923, 1940, 1957, 1974, 1991, 2008,2025, 2042, 2059, 2093, 2110, 2127, 2161, 2178, 2195, 2212, 2229, 2246,2263, 2280, 2314, 2331, 2348, 2365, 2382, 2399, 2416, 2433, 2450, 2467,2484, 2501, 2518, 2535, 2552, 2569, 2586, 2603, 2620, 2637, 2654, 2671,2688, 2705, 2722, 2739, 2773, 2790, 2824, 2841, 2858, 2926, 2943, 2960,3011, 3028, 3062, 3079, 3096, 3113, 3130, 3147, 3164, 3181, 3198, 3232,3249, 3266, and 3283 and/or a recombinant, purified, and/or isolatedpolypeptide selected from the group consisting of a polypeptide havingat least 90% sequence identity to at least one of the sequences of SEQID Nos. 1, 18, 35, 52, 69, 86, 103, 120, 137, 154, 171, 188, 205, 222,239, 256, 273, 290, 307, 324, 358, 375, 409, 426, 443, 511, 528, 545,596, 613, 647, 664, 681, 698, 715, 732, 749, 766, 783, 817, 834, 851,885, 902, 819, 936, 953, 970, 987, 1004, 1038, 1055, 1072, 1089, 1106,1123, 1140, 1157, 1174, 1191, 1208, 1225, 1242, 1259, 1276, 1293, 1310,1327, 1344, 1361, 1378, 1395, 1412, 1429, 1446, 1463, 1480, 1497, 1514,1531, 1548, 1565, 1582, 1599, 1633, 1650, 1684, 1701, 1718, 1786, 1803,1820, 1871, 1888, 1922, 1939, 1956, 1973, 1990, 2007, 2024, 2041, 2058,2092, 2109, 2126, 2160, 2177, 2094, 2211, 2228, 2245, 2262, 2279, 2313,2330, 2347, 2364, 2381, 2398, 2415, 2432, 2449, 2466, 2483, 2500, 2517,2534, 2551, 2568, 2585, 2602, 2619, 2636, 2653, 2670, 2687, 2704, 2721,2738, 2772, 2789, 2823, 2840, 2857, 2925, 2942, 2959, 3010, 3027, 3061,3078, 3095, 3112, 3129, 3146, 3163, 3180, 3197, 3231, 3248, 3265, and3282 to a nucleotide sequence with which recombination is desired.

Further embodiments of methods include placing a cell producing orencoding a recombinant, purified, and/or isolated nucleotide sequencecomprising a nucleotide sequence selected from the group consisting of anucleotide sequences having at least 90% sequence identity to at leastone of the sequences of SEQ ID Nos. 2, 19, 36, 53, 70, 87, 104, 121,138, 155, 172, 189, 206, 223, 240, 257, 274, 291, 308, 325, 359, 376,410, 427, 444, 512, 529, 546, 597, 614, 648, 665, 682, 699, 716, 733,750, 767, 784, 818, 835, 852, 886, 903, 920, 937, 954, 971, 988, 1005,1039, 1056, 1073, 1090, 1107, 1124, 1141, 1158, 1175, 1192, 1209, 1226,1243, 1260, 1277, 1294, 1311, 1328, 1345, 1362, 1379, 1396, 1413, 1430,1447, 1464, 1481, 1498, 1515, 1532, 1549, 1566, 1583, 1600, 1634, 1651,1685, 1702, 1719, 1787, 1804, 1821, 1872, 1889, 1923, 1940, 1957, 1974,1991, 2008, 2025, 2042, 2059, 2093, 2110, 2127, 2161, 2178, 2195, 2212,2229, 2246, 2263, 2280, 2314, 2331, 2348, 2365, 2382, 2399, 2416, 2433,2450, 2467, 2484, 2501, 2518, 2535, 2552, 2569, 2586, 2603, 2620, 2637,2654, 2671, 2688, 2705, 2722, 2739, 2773, 2790, 2824, 2841, 2858, 2926,2943, 2960, 3011, 3028, 3062, 3079, 3096, 3113, 3130, 3147, 3164, 3181,3198, 3232, 3249, 3266, and 3283 and/or a recombinant, purified, and/orisolated polypeptide selected from the group consisting of a polypeptidehaving at least 90% sequence identity to at least one of the sequencesof SEQ ID Nos. 1, 18, 35, 52, 69, 86, 103, 120, 137, 154, 171, 188, 205,222, 239, 256, 273, 290, 307, 324, 358, 375, 409, 426, 443, 511, 528,545, 596, 613, 647, 664, 681, 698, 715, 732, 749, 766, 783, 817, 834,851, 885, 902, 819, 936, 953, 970, 987, 1004, 1038, 1055, 1072, 1089,1106, 1123, 1140, 1157, 1174, 1191, 1208, 1225, 1242, 1259, 1276, 1293,1310, 1327, 1344, 1361, 1378, 1395, 1412, 1429, 1446, 1463, 1480, 1497,1514, 1531, 1548, 1565, 1582, 1599, 1633, 1650, 1684, 1701, 1718, 1786,1803, 1820, 1871, 1888, 1922, 1939, 1956, 1973, 1990, 2007, 2024, 2041,2058, 2092, 2109, 2126, 2160, 2177, 2094, 2211, 2228, 2245, 2262, 2279,2313, 2330, 2347, 2364, 2381, 2398, 2415, 2432, 2449, 2466, 2483, 2500,2517, 2534, 2551, 2568, 2585, 2602, 2619, 2636, 2653, 2670, 2687, 2704,2721, 2738, 2772, 2789, 2823, 2840, 2857, 2925, 2942, 2959, 3010, 3027,3061, 3078, 3095, 3112, 3129, 3146, 3163, 3180, 3197, 3231, 3248, 3265,and 3282 in a environment comprising temperatures at or above about 25,30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, and/or 95 degreesCelsius and/or a pH at, below, and/or above 8, 7, 6, 5, 4, 3, 2, 1,and/or 0.

These and other aspects of the invention will become apparent to theskilled artisan in view of the teachings contained herein.

BRIEF DESCRIPTION OF THE SEVERAL VIEWS OF THE DRAWINGS

FIG. 1 depicts a sequence alignment between SEQ ID NO: 1 (RAAC03697) andref|ZP_(—)01916690.1|, ref|YP_(—)308280.1|, ref|XP_(—)001525241.1|,gb|AAY21825.1|, and ref|XP_(—)001743680.1| (SEQ ID Nos: 3-7respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 2 depicts a sequence alignment between SEQ ID NO: 18 (RAAC02297)and ref|ZP_(—)01171092.1|, ref|YP_(—)429214.1|, gb|EAZ41188.1|,gb|AAR38445.1|, and ref|ZP_(—)01774730.1| (SEQ ID Nos: 20-24respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 3 depicts a sequence alignment between SEQ ID NO: 35 (RAAC02298)and ref|ZP_(—)01575699.1|, ref|ZP_(—)01171091.1|, ref|ZP_(—)02598168.1|,ref|ZP_(—)01900573.1|, and ref|ZP_(—)01301851.1| (SEQ ID Nos: 37-41respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 4 depicts a sequence alignment between SEQ ID NO: 52 (RAAC02299)and ref|ZP_(—)01171090.1|, ref|ZP_(—)02598167.1|, ref|ZP_(—)01575700.1|,ref|ZP_(—)02849387.1|, and ref|NP_(—)627754.1| (SEQ ID Nos: 54-58respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 5 depicts a sequence alignment between SEQ ID NO: 69 (RAAC02300)and ref|ZP_(—)02598166.1|, ref|ZP_(—)01171089.1|, ref|ZP_(—)02849386.1|,ref|YP_(—)159112.1|, and ref|ZP_(—)02007550.1| (SEQ ID Nos: 71-75respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 6 depicts a sequence alignment between SEQ ID NO: 86 (RAAC02301)and ref|ZP_(—)02756760.1|, ref|ZP_(—)01171088.1|, emb|CAE47790.1|,emb|CAE47778.1|, and ref|NP_(—)967133.1| (SEQ ID Nos: 88-92respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 7 depicts a sequence alignment between SEQ ID NO: 103 (RAAC02302)and ref|ZP_(—)01171087.1|, ref|YP_(—)061819.1|, emb|CAJ49597.1|,ref|YP_(—)158155.1|, and ref|NP_(—)779769.1| (SEQ ID Nos: 105-109respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 8 depicts a sequence alignment between SEQ ID NO: 120 (RAAC02303)and ref|YP_(—)001371728.1|, ref|YP_(—)001235767.1|, ref|YP_(—)674884.1|,ref|NP_(—)046584.1|, and ref|ZP_(—)00630666.11 (SEQ ID Nos: 122-126respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 9 depicts a sequence alignment between SEQ ID NO: 137 (RAAC02304)and ref|YP_(—)793245.1|, ref|YP_(—)386759.1|, ref|YP_(—)868126.1|,ref|NP_(—)253469.1|, and ref|ZP_(—)01591801.1| (SEQ ID Nos: 139-143respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 10 depicts a sequence alignment between SEQ ID NO: 154 (RAAC02305)and ref|YP_(—)645800.1|, ref|ZP_(—)01697403.1|, ref|NP_(—)111721.1|,dbj|BAB60367.1|, and ref|YP_(—)950098.11 (SEQ ID Nos: 156-160respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 11 depicts a sequence alignment between SEQ ID NO: 171 (RAAC02306)and gb|EAU91762.1|, ref|ZP_(—)01035289.1|, ref|ZP_(—)01076306.1|,ref|YP_(—)173223.1|, and ref|ZP_(—)01500882.1| (SEQ ID Nos: 173-177respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 12 RAAC02306 RAAC02306 RAAC02306 depicts a sequence alignmentbetween SEQ ID NO: 188 (RAAC02289) and ref|YP_(—)517477.1|,ref|YP_(—)001568284.1|, ref|YP_(—)503850.1|, ref|NP_(—)783815.1|, andref|YP_(—)149134.1| (SEQ ID Nos: 190-194 respectively). Amino acidsconserved among all sequences are indicated by a “*” and generallyconserved amino acids are indicated by a “:”.

FIG. 13 depicts a sequence alignment between SEQ ID NO: 205 (RAAC02307)and ref|XP_(—)001317319.1|, ref|YP_(—)303751.1|, ref|ZP_(—)01222568.1|,ref|XP_(—)001191064.1|, and ref|ZP_(—)01697132.1| (SEQ ID Nos: 207-211respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 14 depicts a sequence alignment between SEQ ID NO: 222 (RAAC02309)and ref|NP_(—)623664.1|, ref|ZP_(—)02755290.1|, ref|YP_(—)001662357.1|,ref|YP_(—)001666189.1|, and ref|YP_(—)001181426.1| (SEQ ID Nos: 224-228respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 15 depicts a sequence alignment between SEQ ID NO: 239 (RAAC02310)and ref|NP_(—)735797.1|, ref|XP_(—)001247966.1|,gb|AAG38042.1|AF295925_(—)7, ref|ZP_(—)00874806.1|, and gb|ABV55445.1|(SEQ ID Nos: 241-245 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 16 depicts a sequence alignment between SEQ ID NO: 256 (RAAC02311)and ref|YP_(—)001205829.1|, ref|YP_(—)001240062.1|, ref|YP_(—)359336.1|,ref|YP_(—)001451893.1|, and ref|YP_(—)466026.1| (SEQ ID Nos: 258-262respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 17 depicts a sequence alignment between SEQ ID NO: 273 (RAAC02312)and ref|YP_(—)001506532.1|, ref|NP_(—)279998.1|, ref|ZP_(—)02850831.1|,ref|ZP_(—)02248080.1|, and ref|XP_(—)001637270.1| (SEQ ID Nos: 275-279respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 18 depicts a sequence alignment between SEQ ID NO: 290 (RAAC02313)and ref|YP_(—)245669.1|, ref|ZP_(—)02369868.1|, ref|YP_(—)438666.1|,ref|NP_(—)695275.1|, and emb|CAB06069.2| (SEQ ID Nos: 292-296respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 19 depicts a sequence alignment between SEQ ID NO: 307 (RAAC02314)and ref|XP_(—)503244.1|, ref|NP_(—)218466.1|, ref|ZP_(—)01058859.1|,ref|NP_(—)126488.1|, and ref|YP_(—)754274.1| (SEQ ID Nos: 309-313respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 20 depicts a sequence alignment between SEQ ID NO: 324 (RAAC02315)and ref|ZP_(—)02854145.1|, ref|YP_(—)145847.1|, ref|YP_(—)536482.1|,ref|YP_(—)799230.1|, and ref|NP_(—)714527.1| (SEQ ID Nos: 326-330respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 21 depicts a sequence alignment between SEQ ID NO: 341 (RAAC02316)and ref|ZP_(—)02180762.1|, ref|ZP_(—)02077766.1|, ref|ZP_(—)01893908.1|,ref|XP^(—)001444409.1|, and ref|CP_(—)624126.2| (SEQ ID Nos: 343-347respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 22 depicts a sequence alignment between SEQ ID NO: 358 (RAAC02290)and emb|CAJ50746.1|, gb|AAM28266.1|, ref|XP_(—)816394.1|,ref|XP_(—)001585185.1|, and gb|EAU92316.1| (SEQ ID Nos: 360-364respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 23 depicts a sequence alignment between SEQ ID NO: 375 (RAAC02317)and ref|YP_(—)001210712.1|, ref|ZP_(—)02727046.1|,ref|ZP_(—)02758954.1|, ref|ZP_(—)01593342.1|, and ref|ZP_(—)00235902.1|(SEQ ID Nos: 377-381 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 24 depicts a sequence alignment between SEQ ID NO: 409 (RAAC02319)and ref|YP_(—)431168.1|, ref|YP_(—)001212944.1|, ref|YP_(—)754944.1|,ref|YP_(—)754864.1|, and ref|ZP_(—)02171383.1| (SEQ ID Nos: 411-415respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 25 depicts a sequence alignment between SEQ ID NO: 426 (RAAC02320)and ref|YP_(—)431169.1|, ref|YP_(—)754945.1|, ref|YP_(—)754863.1|,ref|ZP_(—)01287154.1|, and ref|ZP_(—)01287577.1| (SEQ ID Nos: 428-432respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 26 depicts a sequence alignment between SEQ ID NO: 443 (RAAC02321)and ref|NP_(—)982177.1|, ref|ZP_(—)02595431.1|, ref|NP_(—)927486.1|,ref|YP_(—)001108426.1|, and emb|CAN89659.1| (SEQ ID Nos: 445-449respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 27 depicts a sequence alignment between SEQ ID NO: 460 (RAAC02322)and ref|YP_(—)504284.1|, ref|YP_(—)001046337.1|, ref|ZP_(—)02131576.1|,ref|ZP_(—)01288161.1|, and ref|ZP_(—)02132246.1| (SEQ ID Nos: 462-466respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 28 depicts a sequence alignment between SEQ ID NO: 477 (RAAC02323)and ref|ZP_(—)101287831.1|, ref|YP_(—)076198.1|, ref|YP_(—)446560.1|,ref|NP_(—)634267.1|, and ref|ZP_(—)01106621.1| (SEQ ID Nos: 479-483respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 29 depicts a sequence alignment between SEQ ID NO: 494 (RAAC02324)and ref|ZP_(—)02595423.1|, ref|NP_(—)982173.1|, ref|ZP_(—)02367476.1|,ref|YP_(—)001616264.1|, and ref|YP_(—)827637.1| (SEQ ID Nos: 496-500respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 30 depicts a sequence alignment between SEQ ID NO: 511 (RAAC02326)and ref|NP_(—)982172.1|, ref|ZP_(—)01872101.1|, ref|NP_(—)922949.1|,emb|CAO48005.1|, and sp|P08995|NO26_SOYBN (SEQ ID Nos: 513-517respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 31 depicts a sequence alignment between SEQ ID NO: 528 (RAAC02327)and ref|NP_(—)982171.1|, ref|ZP_(—)01090358.1|, ref|ZP_(—)01856486.1|,ref|ZP_(—)02736297.1|, and ref|ZP_(—)01311632.1| (SEQ ID Nos: 530-534respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 32 depicts a sequence alignment between SEQ ID NO: 545 (RAAC02328)and ref|NP_(—)982170.1|, ref|XP_(—)955124.1|, ref|XP_(—)763458.1|,ref|XP_(—)845342.1|, and ref|XP_(—)666904.1| (SEQ ID Nos: 547-551respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 33 depicts a sequence alignment between SEQ ID NO: 562 (RAAC02332)and ref|YP_(—)079109.1|, ref|ZP_(—)01858609.1|, gb|AAV70501.1|,ref|YP_(—)001319533.1|, and ref|ZP_(—)00539168.1| (SEQ ID Nos: 564-568respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 34 depicts a sequence alignment between SEQ ID NO: 596 (RAAC02334)and ref|YP_(—)001485227.1|, ref|YP_(—)001358015.1|,ref|ZP_(—)02013298.1|, ref|ZP_(—)02178354.1|, and ref|YP_(—)001356736.1|(SEQ ID Nos: 598-602 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 35 depicts a sequence alignment between SEQ ID NO: 613 (RAAC02335)and dbj|BAF91394.11, gb|AAL17690.1|, dbj|BAF91409.1|,ref|NP_(—)001058416.1|, and dbj|BAD45624.1| (SEQ ID Nos: 615-619respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 36 depicts a sequence alignment between SEQ ID NO: 630 (RAAC02336)and ref|YP_(—)517477.1|, ref|YP_(—)517489.1|, ref|ZP_(—)01370335.1|,gb|ACA46983.1|, and ref|YP_(—)001568284.1| (SEQ ID Nos: 632-636respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 37 depicts a sequence alignment between SEQ ID NO: 647 (RAAC02292)and gb|AAB91591.1|, ref|YP_(—)001422657.1|, ref|NP_(—)391247.1|,ref|YP_(—)093160.1|, and ref|NP_(—)391246.1| (SEQ ID Nos: 649-653respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 38 depicts a sequence alignment between SEQ ID NO: 664 (RAAC02337)and gb|ACA41259.1|, ref|ZP_(—)02626811.1|, ref|NP_(—)664934.1|,ref|YP_(—)195796.1|, and ref|NP_(—)817052.1| (SEQ ID Nos: 666-670respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 39 depicts a sequence alignment between SEQ ID NO: 681 (RAAC02338)and gb|ACA41261.1|, ref|ZP_(—)02626812.1|, ref|NP_(—)664935.1|,ref|XP_(—)001701427.1|, and sp|Q2M3V2|ANR43_HUMAN (SEQ ID Nos: 683-687respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 40 depicts a sequence alignment between SEQ ID NO: 698 (RAAC02339)and gb|AAU83457.1|, emb|CAJ70907.1|, ref|ZP_(—)00514953.1|,ref|YP_(—)322920.1|, and ref|YP_(—)183482.1| (SEQ ID Nos: 700-704respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 41 depicts a sequence alignment between SEQ ID NO: 715 (RAAC02340)and ref|ZP_(—)01731985.1|, ref|ZP_(—)02429891.1|, ref|ZP_(—)02432977.1|,ref|ZP_(—)02085861.1|, and ref|ZP_(—)02207628.1| (SEQ ID Nos: 717-721respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 42 depicts a sequence alignment between SEQ ID NO: 732 (RAAC02341)and ref|ZP02596024.1|, ref|NP_(—)150014.1|, ref|ZP_(—)00231288.1|,ref|YP_(—)001213007.1|, and ref|YP_(—)001113884.1| (SEQ ID Nos: 734-738respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 43 depicts a sequence alignment between SEQ ID NO: 749 (RAAC02342)and ref|NP_(—)783868.1|, ref|YP_(—)805310.1|, ref|ZP_(—)01273840.1|,ref|YP_(—)803555.1|, and ref|YP_(—)001270615.1| (SEQ ID Nos: 751-755respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 44 depicts a sequence alignment between SEQ ID NO: 766 (RAAC02293)and ref|ZP_(—)01171099.1|, ref|ZP_(—)02598174.1|, ref|YP 429218.1|,ref|YP_(—)001112194.1|, and ref|YP_(—)001112320.1| (SEQ ID Nos: 768-772respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 45 depicts a sequence alignment between SEQ ID NO: 783 (RAAC02346)and ref|NP_(—)623604.1|, ref|ZP_(—)00235040.1|, ref|ZP_(—)02327783.1|,ref|ZP_(—)02082031.1|, and ref|ZP_(—)02846176.1| (SEQ ID Nos: 785-789respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 46 depicts a sequence alignment between SEQ ID NO: 800 (RAAC02347)and ref|XP_(—)001383704.2|, ref|XP_(—)001664270.1|, ref|YP_(—)113896.1|,ref|XP_(—)761114.1|, and ref|XP_(—)001015776.2| (SEQ ID Nos: 802-806respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 47 depicts a sequence alignment between SEQ ID NO: 817 (RAAC03510)and gb|ACA42330.1|, ref|ZP_(—)02605128.1|, ref|ZP_(—)02586769.1|,ref|YP_(—)001049647.1|, and ref|YP_(—)913053.1| (SEQ ID Nos: 819-823respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 48 depicts a sequence alignment between SEQ ID NO: 834 (RAAC02348)and ref|XP_(—)414088.2|, ref|ZP_(—)01171110.1|, gb|AAF98351.1|,ref|NP_(—)607899.1|, and ref|NP_(—)269831.1| (SEQ ID Nos: 836-840respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 49 depicts a sequence alignment between SEQ ID NO: 851 (RAAC02349)and ref|YP_(—)001038860.1|, ref|YP_(—)754926.1|, ref|YP_(—)001396671.1|,ref|YP_(—)001254879.1|, and ref|YP_(—)001396310.1| (SEQ ID Nos: 853-857respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 50 depicts a sequence alignment between SEQ ID NO: 868 (RAAC03270)and ref|YP_(—)001655174.1|, emb|CAJ73677.1|, emb|CAJ73386.1|,emb|CAJ74660.1|, and ref|ZP_(—)00516046.1| (SEQ ID Nos: 870-874respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 51 depicts a sequence alignment between SEQ ID NO: 885 (RAAC03271)and sp|P26545|VE2_HPV5B, ref|XP_(—)001371550.1|, sp|P36786|VE2_(—HPV)19,ref|NP_(—)041368.1|, and ref|XP_(—)001131003.1| (SEQ ID Nos: 887-891respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 52 depicts a sequence alignment between SEQ ID NO: 902 (RAAC02294)and ref|YP_(—)941474.2|, gb|AAZ42391.1|, ref|ZP_(—)01171098.1|,prf∥112123261AD, and ref|NP_(—)498368.2| (SEQ ID Nos: 904-908respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 53 depicts a sequence alignment between SEQ ID NO: 919 (RAAC02353)and ref|YP_(—)430185.1|, ref|YP_(—)001233893.1|, emb|CA081523.1|,ref|YP_(—)001603689.1|, and ref|YP_(—)719187.1| (SEQ ID Nos: 921-925respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 54 depicts a sequence alignment between SEQ ID NO: 936 (RAAC02354)and ref|YP_(—)001438903.1|, ref|YP_(—)001251565.1|, ref|YP_(—)126374.1|,ref|YP_(—)123382.1|, and ref|YP_(—)001251158.1| (SEQ ID Nos: 938-942respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 55 depicts a sequence alignment between SEQ ID NO: 953 (RAAC02355)and ref|ZP_(—)02758276.1|, gb|ACA42232.1|, ref|ZP_(—)02602342.1|,ref|ZP_(—)02597242.1|, and ref|ZP_(—)02524501.1| (SEQ ID Nos: 955-959respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 56 depicts a sequence alignment between SEQ ID NO: 970 (RAAC02356)and ref|NP_(—)347717.1|, ref|YP_(—)423535.1|, ref|ZP_(—)01860459.1|,ref|YP_(—)429187.1|, and ref|YP_(—)645289.1| (SEQ ID Nos: 972-976respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 57 depicts a sequence alignment between SEQ ID NO: 987 (RAAC02357)and gb|EAY58379.1|, ref|ZP_(—)01505670.1|, ref|NP_(—)856790.1|,ref|NP_(—)217638.1|, and ref|ZP_(—)00876805.1| (SEQ ID Nos: 989-993respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 58 depicts a sequence alignment between SEQ ID NO: 1004 (RAAC02358)and ref|YP_(—)024839.1|, ref|YP_(—)001456771.1|, ref|YP_(—)655149.1|,ref|XP_(—)748956.1|, and ref|NP_(—)943831.1| (SEQ ID Nos: 1006-1010respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 59 depicts a sequence alignment between SEQ ID NO: 1038 (RAAC02361)and ref|ZP_(—)01966753.1|, ref|YP_(—)001086797.1|,ref|ZP_(—)01805266.1|, ref|YP_(—)001681547.1|, and ref|ZP_(—)01188985.1|(SEQ ID Nos: 1040-1044 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 60 depicts a sequence alignment between SEQ ID NO: 1055 (RAAC02362)and ref|YP_(—)157691.1|, ref|ZP_(—)02509777.1|, ref|ZP_(—)02485527.1|,ref|ZP_(—)02406784.1|, and ref|YP_(—)335221.1| (SEQ ID Nos: 1057-1061respectively) 1055 in Table 1. Amino acids conserved among all sequencesare indicated by a “*” and generally conserved amino acids are indicatedby a “:”.

FIG. 61 depicts a sequence alignment between SEQ ID NO: 1072 (RAAC02363)and ref|YP_(—)001681084.1|, ref|YP_(—)177318.1|, ref|ZP_(—)02329650.1|,ref|NP_(—)243607.1|, and ref|YP_(—)001421775.1| (SEQ ID Nos: 1074-1078respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 62 depicts a sequence alignment between SEQ ID NO: 1089 (RAAC02364)and ref|YP_(—)079689.1|, ref|ZP_(—)02329649.1|, ref|ZP_(—)01860132.1|,ref|ZP_(—)171904.1|, and ref|YP_(—)001487332.1| (SEQ ID Nos: 1091-1095respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 63 depicts a sequence alignment between SEQ ID NO: 1106 (RAAC02366)and ref|ZP_(—)01614696.1|, gb|ABH06559.1|, gb|AAB95339.1|,ref|YP_(—)161675.1|, and ref|YP_(—)001202661.1| (SEQ ID Nos: 1108-1112respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 64 depicts a sequence alignment between SEQ ID NO: 1123 (RAAC02367)and ref|ZP_(—)02093159.1|, ref|ZP_(—)02026447.1|,ref|YP_(—)001127515.1|, ref|ZP_(—)02091713.1|, and ref|ZP_(—)02423704.1|(SEQ ID Nos: 1125-1129 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 65 depicts a sequence alignment between SEQ ID NO: 1140 (RAAC02370)and ref|ZP_(—)00960984.1|, ref|ZP_(—)01035782.1|, ref|ZP_(—)01903846.1|,ref|ZP_(—)880414.1|, and ref|ZP_(—)01742943.1| (SEQ ID Nos: 1142-1146respectively). Amino acids conserved among all sequences are indicatedby a “*” and gene conserved amino acids are indicated by a “:”.

FIG. 66 depicts a sequence alignment between SEQ ID NO: 1157 (RAAC02371)and ref|ZP_(—)02756730.1|, ref|ZP_(—)02833143.1|, ref|YP_(—)521772.1|,ref|ZP_(—)01551668.1|, and ref|YP_(—)887014.1| (SEQ ID Nos: 1159-1163respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 67 depicts a sequence alignment between SEQ ID NO: 1174 (RAAC02372)and sp|Q7ZXB1|MCM7B_XENLA, ref|YP_(—)324842.1|, ref|NP_(—)486002.1|,ref|XP_(—)660834.1|, and ref|ZP_(—)01733540.1| (SEQ ID Nos: 1176-1180respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 68 depicts a sequence alignment between SEQ ID NO: 1191 (RAAC02296)and ref|XP_(—)001563017.1|, ref|XP_(—)001615133.1|,ref|YP_(—)001236354.1|, ref|ZP_(—)0177649.1|, and ref|ZP_(—)01565636.1|(SEQ ID Nos: 1193-1197 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 69 depicts a sequence alignment between SEQ ID NO: 1208 (RAAC02373)and ref|YP_(—)001376930.1|, gb|AAW81277.1|, ref|ZP_(—)02852259.1|,ref|YP_(—)001642790.1|, and ref|YP_(—)001312077.1| (SEQ ID Nos:1210-1214 respectively). Amino acids conserved among all sequences areindicated by a “*” and generally conserved amino acids are indicated bya “:”.

FIG. 70 depicts a sequence alignment between SEQ ID NO: 1225 (RAAC02374)and ref|XP_(—)001467069.1|, ref|YP_(—)001376929.1|,ref|YP_(—)001208199.1|, gb|EAU86007.1|, and ref|YP_(—)946581.1| (SEQ IDNos: 1227-1231 respectively). Amino acids conserved among all sequencesare indicated by a “*” and generally conserved amino acids are indicatedby a “:”.

FIG. 71 depicts a sequence alignment between SEQ ID NO: 1242 (RAAC02375)and ref|XP_(—)975359.1|, ref|XP_(—)001315633.1|, ref|ZP_(—)02840410.1|,ref|ZP_(—)01467536.1|, and ref|YP_(—)001016790.1| (SEQ ID Nos: 1244-1248respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 72 depicts a sequence alignment between SEQ ID NO: 1259 (RAAC03273)and ref|YP_(—)502758.1|, gb|EAU81483.1|, ref|ZP_(—)01091610.1|,ref|YP_(—)944003.1|, and ref|YP_(—)462360.1| (SEQ ID Nos: 1261-1265respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 73 depicts a sequence alignment between SEQ ID NO: 1276 (RAAC02967)and ref|YP_(—)074959.1|, ref|YP_(—)594046.1|, ref|ZP_(—)01846154.1|,sp|Q45618|TRA6_BACST, and ref|YP_(—)828009.1| (SEQ ID Nos: 1278-1282respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 74 depicts a sequence alignment between SEQ ID NO: 1293 (RAAC03589)and ref|YP_(—)146741.1|, ref|YP_(—)148969.1|, ref|YP_(—)001126171.1|,ref|ZP_(—)02130848.1|, and ref|YP_(—)146154.1| (SEQ ID Nos: 1295-1299respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 75 depicts a sequence alignment between SEQ ID NO: 1310 (RAAC03695)and ref|ZP101665148.1|, ref|YP_(—)001111903.1|, ref|YP_(—)752864.1|,ref|YP_(—)753434.1|, and ref|YP_(—)753226.1| (SEQ ID Nos: 1312-1316respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 76 depicts a sequence alignment between SEQ ID NO: 1327 (RAAC02318)and ref|YP_(—)754943.1|, ref|YP_(—)754865.1|, ref|YP 431166.1|,ref|ZP_(—)02171171.1|, and ref|YP_(—)519650.1| (SEQ ID Nos: 1329-1333respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 77 depicts a sequence alignment between SEQ ID NO: 1344 (RAAC02319)and ref|YP_(—)431168.1|, ref|YP_(—)001212944.1|, ref|YP_(—)754944.1|,ref|YP_(—)754864.1|, and ref|ZP_(—)02171383.1| (SEQ ID Nos: 1346-1350respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 78 depicts a sequence alignment between SEQ ID NO: 1361 (RAAC02333)and ref|ZP02734990.1|, ref|ZP_(—)01265219.1|, ref|YP_(—)266430.1|,ref|YP_(—)008142.1|, and ref|YP_(—)713924.1| (SEQ ID Nos: 1363-1367respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 79 depicts a sequence alignment between SEQ ID NO: 1378 (RAAC03703)and ref|YP_(—)001036724.1|, ref|YP_(—)001039064.1|,ref|YP_(—)001039349.1|, ref|Y_(—)076118.1|, and ref|YP_(—)074958.1|X(SEQ ID Nos: 1380-1384 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 80 depicts a sequence alignment between SEQ ID NO: 1395 (RAAC03568)and ref|YP_(—)001039349.1|, ref|YP_(—)001036724.1|,ref|YP_(—)001039064.1|, ref|YP_(—)076118.1|, and ref|YP_(—)074958.1|X(SEQ ID Nos: 1397-1401 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 81 depicts a sequence alignment between SEQ ID NO: 1412 (RAAC03707)and ref|YP_(—)519534.1|, ref|ZP_(—)01370818.1|, ref|ZP_(—)01372264.1|,ref|YP_(—)516922.1|, and ref|YP_(—)519084.1|X (SEQ ID Nos: 1414-1418respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 82 depicts a sequence alignment between SEQ ID NO: 1429 (RAAC03173)and ref|YP_(—)431168.1|, ref|YP_(—)001212944.1|, ref|YP_(—)754944.1|,ref|YP_(—)754864.1|, and ref|ZP_(—)02171383.1|X (SEQ ID Nos: 1431-1435respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 83 depicts a sequence alignment between SEQ ID NO: 1446 (RAAC02966)and ref|YP_(—)074959.1|, ref|NP_(—)634718.1|, ref|NP_(—)616807.1|,gb|AAR99616.1|, and sp|Q45618|TRA6_BACSTX (SEQ ID Nos: 1448-1452respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 84 depicts a sequence alignment between SEQ ID NO: 1463 (RAAC00757)and ref|YP_(—)430545.1|, ref|YP_(—)148444.1|, ref|NP_(—)980798.1|,ref|YP_(—)001211577.1|, and sp|Q9KDI8|RUVB_BACHDX (SEQ ID Nos: 1465-1469respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 85 depicts a sequence alignment between SEQ ID NO: 1480 (RAAC00756)and ref|YP_(—)234498.1|, ref|NP_(—)793742.1|, ref|YP_(—)275913.1|,ref|ZP_(—)00991066.1|, and ref|YP_(—)001186768.1|X (SEQ ID Nos:1482-1486 respectively). Amino acids conserved among all sequences areindicated by a “*” and generally conserved amino acids are indicated bya “:”.

FIG. 86 depicts a sequence alignment between SEQ ID NO: 1497 (RAAC00755)and ref|YP_(—)644098.1|, ref|ZP_(—)02848139.1|, ref|YP_(—)518701.1|,ref|YP_(—)074988.1|, and ref|YP_(—)001180347.1|X (SEQ ID Nos: 1499-1503respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 87 depicts a sequence alignment between SEQ ID NO: 1514 (RAAC01468)and ref|YP_(—)146341.1|, ref|YP_(—)001513188.1|, ref|ZP_(—)02261478.1|,ref|YP_(—)001373830.1|, and ref|ZP_(—)02257063.1|X (SEQ ID Nos:1516-1520 respectively). Amino acids conserved among all sequences areindicated by a “*” and generally conserved amino acids are indicated bya “:”.

FIG. 88 depicts a sequence alignment between SEQ ID NO: 1531 (RAAC03178)and ref|ZP_(—)01665148.1|, ref|YP_(—)752864.1|, ref|YP_(—)001111903.1|,ref|YP_(—)753434.1|, and ref|YP_(—)753226.1|X (SEQ ID Nos: 1533-1537respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 89 depicts a sequence alignment between SEQ ID NO: 1548 (RAAC01937)and dbj|BAF33373.1|, ref|YP_(—)001126744.1|, gb|AAB52611.1|,gb|ABM97416.1|, and sp|P52026|DPO1_BACSTX (SEQ ID Nos: 1550-1554respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 90 depicts a sequence alignment between SEQ ID NO: 1565 (RAAC01372)and ref|YP_(—)001681573.1|, ref|ZP_(—)00651175.1|, ref|YP_(—)473713.1|,gb|ACA11657.1|, and ref|YP_(—)475384.1|X (SEQ ID Nos: 1567-1571respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 91 depicts a sequence alignment between SEQ ID NO: 1582 (RAAC00062)and ref|ZP_(—)01695687.1|, ref|ZP01695971.1|, ref|ZP_(—)01695982.1|,ref|ZP_(—)01695655.1|, and ref|YP_(—)430569.1|X (SEQ ID Nos: 1584-1588respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 92 depicts a sequence alignment between SEQ ID NO: 1599 (RAAC02377)and ref|YP_(—)752864.1|, ref|ZP_(—)01665148.1|, ref|YP_(—)001111903.1|,ref|YP_(—)754667.1|, and ref|YP_(—)753226.1|X (SEQ ID Nos: 1601-1605respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 93 depicts a sequence alignment between SEQ ID NO: 1633 (RAAC03117)and ref|YP_(—)754865.1|, ref|YP_(—)754943.1|, ref|YP_(—)431166.1|,ref|ZP02171171.1|, and ref|YP_(—)001318017.1|X (SEQ ID Nos: 1635-1639respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 94 depicts a sequence alignment between SEQ ID NO: 1650 (RAAC00037)and ref|YP_(—)001514390.1|, ref|NP_(—)780819.1|, ref|YP_(—)001663996.1|,ref|YP_(—)699899.1|, and ref|YP_(—)077153.1|X (SEQ ID Nos: 1652-1656respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 95 depicts a sequence alignment between SEQ ID NO: 1667 (RAAC00054)and ref|NP_(—)387885.1|, ref|YP_(—)803557.1|, ref|YP_(—)077286.1|,ref|ZP_(—)02326643.1|, and ref|YP_(—)001419683.1|X (SEQ ID Nos:1669-1673 respectively). Amino acids conserved among all sequences areindicated by a “*” and generally conserved amino acids are indicated bya “:”.

FIG. 96 depicts a sequence alignment between SEQ ID NO: 1684 (RAAC03102)and ref|YP_(—)074959.1|, gb|AAR99616.1|, sp|Q45618|TRA6_BACST,ref|YP_(—)075129.1|, and ref|YP_(—)359963.1|X (SEQ ID Nos: 1686-1690respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 97 depicts a sequence alignment between SEQ ID NO: 1701 (RAAC03103)and ref|YP_(—)074959.1|, ref|YP_(—)594046.1|, ref|ZP_(—)02563000.1|,ref|ZP_(—)02516401.1|, and ref|ZP_(—)02335796.1|X (SEQ ID Nos: 1703-1707respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 98 depicts a sequence alignment between SEQ ID NO: 1718 (RAAC03341)and ref|YP_(—)076075.1|, ref|YP_(—)074958.1|, ref|YP_(—)076118.1|,ref|YP_(—)001039349.1|, and ref|YP_(—)001036724.1|X (SEQ ID Nos:1720-1724 respectively). Amino acids conserved among all sequences areindicated by a “*” and generally conserved amino acids are indicated bya “:”.

FIG. 99 depicts a sequence alignment between SEQ ID NO: 1786 (RAAC02852)and ref|YP_(—)001125185.1|, ref|YP_(—)147064.1|, ref|YP_(—)001486753.1|,ref|NP_(—)389494.1|, and ref|YP_(—)091420.1|X (SEQ ID Nos: 1788-1792respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 100 depicts a sequence alignment between SEQ ID NO: 1803(RAAC02854) and ref|YP_(—)001125186.1|, ref|ZP_(—)01775043.1|,ref|YP_(—)175772.1|, ref|NP_(—)243331.1|, and ref|YP_(—)740910.1|X (SEQID Nos: 1805-1809 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 101 depicts a sequence alignment between SEQ ID NO: 1820(RAAC03166) and ref|YP_(—)001211938.1|, dbj|BAD22831.1|,ref|ZP_(—)01666445.1|, ref|ZP_(—)01665334.1|, and gb|EAY56013.1|X (SEQID Nos: 1822-1826 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 102 depicts a sequence alignment between SEQ ID NO: 1854(RAAC02961) and ref|ZP_(—)02330756.1|, ref|ZP_(—)02327778.1|,ref|ZP_(—)02330395.1|, ref|ZP_(—)02327484.1|, and ref|ZP_(—)02326400.1|X(SEQ ID Nos: 1856-1860 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 103 depicts a sequence alignment between SEQ ID NO: 1871(RAAC02202) and ref|YP_(—)146129.1|, sp|O87703|DNLJ_BACST,gb|ABN05294.1|, ref|YP_(—)001124385.1|, and ref|ZP_(—)01169975.1|X (SEQID Nos: 1873-1877 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 104 depicts a sequence alignment between SEQ ID NO: 1888(RAAC03682) and ref|ZP_(—)01695687.1|, ref|ZP_(—)01695971.1|,ref|ZP_(—)01695982.1|, ref|ZP_(—)01695655.1|, and ref|YP_(—)430569.1|X(SEQ ID Nos: 1890-1894 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 105 depicts a sequence alignment between SEQ ID NO: 1922(RAAC03770) and ref|ZP_(—)01695687.1|, ref|ZP_(—)01695971.1|,ref|ZP_(—)695982.1|, ref|ZP_(—)695655.1|, and ref|YP_(—)430569.1|X (SEQID Nos: 1924-1928 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 106 depicts a sequence alignment between SEQ ID NO: 1939(RAAC02738) and ref|NP_(—)842969.1|, ref|YP_(—)430640.1|,ref|ZP_(—)02596019.1|, ref|YP_(—)001666203.1|, andref|YP_(—)001512589.1|X (SEQ ID Nos: 1941-1945 respectively). Aminoacids conserved among all sequences are indicated by a “*” and generallyconserved amino acids are indicated by a “:”.

FIG. 107 depicts a sequence alignment between SEQ ID NO: 1956(RAAC02514) and ref|YP_(—)001114460.1|, ref|YP_(—)001111555.1|,ref|YP_(—)001112147.1|, ref|YP_(—)001111684.1|, andref|YP_(—)001113963.1|X (SEQ ID Nos: 1958-1962 respectively). Aminoacids conserved among all sequences are indicated by a “*” and generallyconserved amino acids are indicated by a “:”.

FIG. 108 depicts a sequence alignment between SEQ ID NO: 1973(RAAC02515) and ref|YP_(—)001111555.1|, ref|YP_(—)001111684.1|,ref|YP_(—)001114460.1|, ref|YP_(—)001113963.1|, and dbj|BAD18231.1|X(SEQ ID Nos: 1975-1979 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 109 depicts a sequence alignment between SEQ ID NO: 1990(RAAC02530) and ref|YP_(—)001212947.1|, ref|YP_(—)431166.1|,ref|YP_(—)754943.1|, ref|YP_(—)754865.1|, and ref|YP_(—)_(—)001318017.1|X (SEQ ID Nos: 1992-1996 respectively). Amino acidsconserved among all sequences are indicated by a “*” and generallyconserved amino acids are indicated by a “:”.

FIG. 110 depicts a sequence alignment between SEQ ID NO: 2007(RAAC02533) and ref|YP_(—)001039349.1|, ref|YP_(—)001036724.1|,ref|YP_(—)001039064.1|, ref|YP_(—)076118.1|, and ref|YP_(—)076073.1|X(SEQ ID Nos: 2009-2013 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 111 depicts a sequence alignment between SEQ ID NO: 2024(RAAC02534) and ref|YP_(—)076118.1|, ref|YP_(—)074958.1|,ref|YP_(—)001039349.1|, ref|YP_(—)001036724.1|, andref|YP_(—)001039064.1|X (SEQ ID Nos: 2026-2030 respectively). Aminoacids conserved among all sequences are indicated by a “*” and generallyconserved amino acids are indicated by a “:”.

FIG. 112 depicts a sequence alignment between SEQ ID NO: 2041(RAAC02562) and ref|YP_(—)148733.1|, ref|YP_(—)146224.1|,ref|ZP_(—)02619781.1|, ref|ZP_(—)02171259.1|, and ref|NP_(—)977687.1|X(SEQ ID Nos: 2043-2047 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 113 depicts a sequence alignment between SEQ ID NO: 2058(RAAC03229) and ref|YP_(—)519534.1|, ref|ZP_(—)01370818.1|,ref|ZP_(—)01372264.1|, ref|YP_(—)516922.1|, and ref|YP_(—)520815.1|X(SEQ ID Nos: 2060-2064 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 114 depicts a sequence alignment between SEQ ID NO: 2075

FIG. 124 depicts a sequence alignment between SEQ ID NO: 2092(RAAC00160) and ref|YP_(—)001125159.1|, ref|ZP_(—)02849289.1|,ref|ZP_(—)02329219.1|, ref|YP_(—)147039.1|, and ref|NP_(—)816723.1|X(SEQ ID Nos: 2094-2098 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 115 depicts a sequence alignment between SEQ ID NO: 2109(RAAC03182) and ref|YP_(—)146741.1|, ref|YP_(—)148969.1|,ref|YP_(—)001126171.1|, ref|ZP_(—)02130848.1|, and ref|YP_(—)146154.1|X(SEQ ID Nos: 2111-2115 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 116 depicts a sequence alignment between SEQ ID NO: 2126(RAAC03163) and ref|YP_(—)148733.1|, ref|YP_(—)146224.1|,ref|ZP_(—)02171259.1|, ref|ZP_(—)02619781.1|, and ref|NP_(—)977687.1|X(SEQ ID Nos: 2128-2132 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 117 depicts a sequence alignment between SEQ ID NO: 2143(RAAC01387) and ref|YP_(—)148733.1|, ref|YP_(—)146224.1|,ref|ZP_(—)02171259.1|, ref|ZP_(—) 02619781.1|, and ref|YP_(—)001396471.1|X (SEQ ID Nos: 2145-2149 respectively). Amino acidsconserved among all sequences are indicated by a “*” and generallyconserved amino acids are indicated by a “:”.

FIG. 118 depicts a sequence alignment between SEQ ID NO: 2160(RAAC03385) and gb|AAL87775.1|AF403183_(—)1286, ref|YP_(—)517288.1|,ref|YP_(—)1517659.1|, ref|ZP01368657.1|, and ref|YP_(—)001212990.1|X(SEQ ID Nos: 2162-2166 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 119 depicts a sequence alignment between SEQ ID NO: 2177(RAAC03398) and ref|YP_(—)001126171.1|, ref|YP_(—)148969.1|,ref|YP_(—)146154.1|, ref|YP_(—)146741.1|, and ref|ZP_(—)02172080.1|X(SEQ ID Nos: 2179-2183 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 120 depicts a sequence alignment between SEQ ID NO: 2194(RAAC03177) and ref|YP_(—)148969.1|, ref|YP_(—)146154.1|,ref|YP_(—)001126171.1|, ref|YP_(—)146741.1|, and ref|ZP_(—)02172080.1|X(SEQ ID Nos: 2196-2200 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 121 depicts a sequence alignment between SEQ ID NO: 2211(RAAC03588) and ref|YP_(—)001036724.1|, ref|YP_(—)001039064.1|,ref|YP_(—)001039349.1|, ref|YP_(—)074105.1|, and ref|YP_(—)076073.1|X(SEQ ID Nos: 2213-2217 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 122 depicts a sequence alignment between SEQ ID NO: 2228(RAAC03818) and ref|YP_(—)074959.1|, ref|NP_(—)634718.1|,gb|AAR99616.1|, sp|Q45618|TRA6_BACST, and ref|NP_(—)616807.1|X (SEQ IDNos: 2230-2234 respectively). Amino acids conserved among all sequencesare indicated by a “*” and generally conserved amino acids are indicatedby a “:”.

FIG. 123 depicts a sequence alignment between SEQ ID NO: 2245(RAAC03819) and ref|YP_(—)594046.1|, ref|YP_(—)074959.1|,ref|YP_(—)359963.1|, ref|YP_(—)361300.1|, and ref|NP_(—)622784.1|X (SEQID Nos: 2247-2251 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 124 depicts a sequence alignment between SEQ ID NO: 2262(RAAC03823) and ref|ZP01695687.1|, ref|ZP_(—)01695971.1|,ref|ZP_(—)01695982.1|, ref|ZP_(—)01695655.1|, and ref|YP_(—)430569.1|X(SEQ ID Nos: 2264-2268 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 125 depicts a sequence alignment between SEQ ID NO: 2279(RAAC01171) and ref|ZP_(—)02330348.1|, ref|ZP_(—)02328298.1|,ref|ZP_(—)02326599.1|, ref|YP_(—)001664428.1|, andref|YP_(—)001664274.1| (SEQ ID Nos: 2281-2285 respectively). Amino acidsconserved among all sequences are indicated by a “*” and generallyconserved amino acids are indicated by a “:”.

FIG. 126 depicts a sequence alignment between SEQ ID NO: 2296(RAAC03825) and ref|YP_(—)519534.1|, ref|ZP_(—)01370818.1|,ref|ZP_(—)01372264.1|, ref|Y_(—)516922.1|, and ref|Y_(—)520815.1| (SEQID Nos: 2298-2302 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 127 depicts a sequence alignment between SEQ ID NO: 2313(RAAC03826) and ref|ZP_(—)02851608.1|, ref|ZP_(—)02326599.1|,ref|YP_(—)001307815.1|, ref|YP_(—)001212943.1|, andref|YP_(—)001113174.1| (SEQ ID Nos: 2315-2319 respectively). Amino acidsconserved among all sequences are indicated by a “*” and generallyconserved amino acids are indicated by a “:”.

FIG. 128 depicts a sequence alignment between SEQ ID NO: 2330(RAAC02717) and ref|ZP 01002160.1|, ref|ZP_(—)01592598.1|,ref|YP_(—)001229345.1|, ref|YP_(—)771767.1|, and ref|YP_(—)245447.1|(SEQ ID Nos: 2332-2336 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 129 depicts a sequence alignment between SEQ ID NO: 2347(RAAC01155) and ref|YP_(—)145872.1|, ref|YP_(—)001124150.1|,ref|ZP_(—)02850412.1|, ref|NP_(—)240902.1|, and ref|NP_(—)466224.1| (SEQID Nos: 2349-2353 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 130 depicts a sequence alignment between SEQ ID NO: 2364(RAAC03145) and dbj|BAA94830.1|, ref|ZP_(—)01666433.1|,ref|YP_(—)001213263.1|, ref|YP_(—)387030.1|, and ref|YP_(—)387237.1|(SEQ ID Nos: 2366-2370 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 131 depicts a sequence alignment between SEQ ID NO: 2381(RAAC03325) and ref|YP_(—)146741.1|, ref|YP_(—)148969.1|,ref|YP_(—)001126171.1|, ref|ZP_(—)02130848.1|, and ref|YP_(—)146154.1|(SEQ ID Nos: 2383-2387 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 132 depicts a sequence alignment between SEQ ID NO: 2398(RAAC03376) and ref|ZP_(—)02851608.1|, ref|ZP_(—)02326599.1|,ref|YP_(—)001664274.1|, ref|YP_(—)001307815.1|, andref|YP_(—)001319448.1| (SEQ ID Nos: 2400-2404 respectively). Amino acidsconserved among all sequences are indicated by a “*” and generallyconserved amino acids are indicated by a “:”.

FIG. 133 depicts a sequence alignment between SEQ ID NO: 2415(RAAC02657) and ref|YP_(—)076090.1|, ref|YP_(—)001516732.1|,ref|YP_(—)001185431.1|, ref|YP_(—)318565.1|, and ref|ZP_(—)01061333.1|(SEQ ID Nos: 2417-2421 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 134 depicts a sequence alignment between SEQ ID NO: 2432(RAAC01373) and ref|YP_(—)001680037.1|, ref|YP_(—)001505049.1|,ref|YP_(—)482514.1|, ref|YP_(—)481774.1|, and ref|NP_(—)215436.1| (SEQID Nos: 2434-2438 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 135 depicts a sequence alignment between SEQ ID NO: 2449(RAAC00337) and ref|YP_(—)001680296.1|, ref|YP_(—)342400.1|,ref|NP_(—)756021.1|, ref|YP_(—)405147.1|, and ref|NP_(—)709160.1| (SEQID Nos: 2451-2455 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 136 depicts a sequence alignment between SEQ ID NO: 2466(RAAC00506) and emb|CAD18993.1|, ref|ZP_(—)00995572.1|,ref|YP_(—)805441.1|, ref|YP_(—)001115956.1|, and ref|ZP_(—)02134324.1|(SEQ ID Nos: 2468-2472 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 137 depicts a sequence alignment between SEQ ID NO: 2483(RAAC00022) and ref|ZP_(—)01695451.1|, ref|NP_(—)391924.1|,ref|YP_(—)001488901.1|, ref|NP_(—)244897.1|, and ref|YP_(—)001423303.1|(SEQ ID Nos: 2485-2489 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 138 depicts a sequence alignment between SEQ ID NO: 2500(RAAC00027) and ref|YP_(—)001213441.1|, ref|NP_(—)244917.1|,ref|YP_(—)001377189.1|, ref|YP_(—)149334.1|, and ref|YP_(—)077145.1|(SEQ ID Nos: 2502-2506 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 139 depicts a sequence alignment between SEQ ID NO: 2517(RAAC01051) and ref|YP_(—)074151.1|, ref|YP_(—)001255315.1|,ref|YP_(—)001392092.1|, gb|ACA43749.1|, and ref|ZP_(—)02619122.1|X (SEQID Nos: 2519-2523 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 140 depicts a sequence alignment between SEQ ID NO: 2534.(RAAC01050) and ref|YP_(—)878438.1|, ref|ZP_(—)02621211.1|,ref|YP_(—)001560315.1|, ref|YP_(—)001392091.1|, andref|YP_(—)001255314.1|X (SEQ ID Nos: 2536-2540 respectively). Aminoacids conserved among all sequences are indicated by a “*” and generallyconserved amino acids are indicated by a “:”.

FIG. 141 depicts a sequence alignment between SEQ ID NO: 2551(RAAC01009) and ref|YP_(—)148023.1|, ref|YP_(—)001126202.1|,ref|YP_(—)001421643.1|, ref|ZP_(—)01697284.1|, and gb|ABN10253.1|X (SEQID Nos: 2553-2557 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 142 depicts a sequence alignment between SEQ ID NO: 2568(RAAC00998) and ref|ZP_(—)02175216.1|, ref|YP 464174.1|,ref|YP_(—)753805.1|, ref|ZP_(—)02321813.1|, and ref|ZP_(—)01575281.1|X(SEQ ID Nos: 2570-2574 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 143 depicts a sequence alignment between SEQ ID NO: 2585(RAAC02359) and ref|NP_(—)832076.1|, ref|YP_(—)001645033.1|,ref|NP_(—)844759.1|, ref|YP_(—)001375058.1|, and ref|YP_(—)535778.1|X(SEQ ID Nos: 2587 FIG. 153-2591 respectively). Amino acids conservedamong all sequences are indicated by a “*” and generally conserved aminoacids are indicated by a “:”.

FIG. 144 depicts a sequence alignment between SEQ ID NO: 2602(RAAC00997) and ref|ZP_(—)02854041.1|, ref|YP_(—)075626.1|,ref|YP_(—)753804.1|, ref|ZP_(—)01667008.1|, and ref|YP_(—)946103.1|X(SEQ ID Nos: 2604-2608 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 145 depicts a sequence alignment between SEQ ID NO: 2619(RAAC02419) and ref|ZP02330146.1|, ref|YP_(—)001210709.1|,ref|ZP_(—)02184702.1|, gb|AAD26564.1|AF124258_(—)1, andref|ZP_(—)01828805.1|X (SEQ ID Nos: 2621-2625 respectively). Amino acidsconserved among all sequences are indicated by a “*” and generallyconserved amino acids are indicated by a “:”.

FIG. 145 depicts a sequence alignment between SEQ ID NO: 2636(RAAC02417) and ref|NP_(—)469419.1|, ref|ZP_(—)02309926.1|,ref|ZP_(—)01926077.1|, ref|ZP_(—)0941236.1|, and ref|YP_(—)001111866.1|X(SEQ ID Nos: 2638-2642 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 147 depicts a sequence alignment between SEQ ID NO: 2653(RAAC03180) and ref|ZP02442523.1|, ref|YP_(—)001664041.1|,ref|YP_(—)001210714.1|, ref|ZP_(—)02589119.1|, and ref|NP_(—)242309.1|X(SEQ ID Nos: 2655-2659 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 148 depicts a sequence alignment between SEQ ID NO: 2670(RAAC03525) and ref|ZP_(—)02442523.1|, ref|YP_(—)001210714.1|,ref|YP_(—)001180871.1|, ref|YP_(—)001662345.1|, andref|ZP_(—)02543721.1|X (SEQ ID Nos: 2672-2676 respectively). Amino acidsconserved among all sequences are indicated by a “*” and generallyconserved amino acids are indicated by a “:”.

FIG. 149 depicts a sequence alignment between SEQ ID NO: 2687(RAAC03224) and ref|YP_(—)006607.1|, ref|YP_(—)001038857.1|,ref|YP_(—)146372.1|, ref|YP_(—)001662865.1|, and ref|ZP_(—)02781438.1|X(SEQ ID Nos: 2689-2693 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 150 depicts a sequence alignment between SEQ ID NO: 2704(RAAC02915) and ref|YP_(—)001211829.1|, ref|ZP_(—)02091210.1|,ref|YP_(—)518738.1|, ref|ZP_(—)02378091.1|, and ref|ZP_(—)02454559.1|X(SEQ ID Nos: 2706-2710 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 151 depicts a sequence alignment between SEQ ID NO: 2721(RAAC02943) and ref|ZP_(—)02850845.1|, ref|NP_(—)243249.1|,emb|CAD56684.1|, ref|YP_(—)079090.1|, and ref|YP_(—)001421272.1|X (SEQID Nos: 2723-2727 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 152 depicts a sequence alignment between SEQ ID NO: 2738(RAAC02234) and ref|NP_(—)242725.1|, ref|YP_(—)175539.1|,ref|ZP_(—)02170975.1|, ref|ZP_(—)01696769.1|, and emb|CAJ73252.1|X (SEQID Nos: 2740-2744 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 153 depicts a sequence alignment between SEQ ID NO: 2755(RAAC01662) and emb|CAK51299.1|, ref|YP_(—)832554.1|, emb|CAI78402.1|,ref|YP_(—)001362100.1|, and ref|YP_(—)712120.1|X (SEQ ID Nos: 2757-2761respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 154 depicts a sequence alignment between SEQ ID NO: 2772(RAAC02171) and ref|ZP_(—)01188667.1|, ref|ZP02620185.1|,ref|YP_(—)001512273.1|, ref|YP_(—)877450.1|, and ref|YP_(—)001275096.1|X(SEQ ID Nos: 2774-2778 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 155 depicts a sequence alignment between SEQ ID NO: 2789(RAAC01696) and sp|P80579|THIO_ALIAC, pdb|1NW2|A, pdb|1NSW|A,pdb|1RQM|A, and ref|YP_(—)703612 .1|X (SEQ ID Nos: 2791-2795respectively). Amino acids conserved among all sequences are indicatedby a “*” and generally conserved amino acids are indicated by a “:”.

FIG. 156 depicts a sequence alignment between SEQ ID NO: 2806(RAAC01724) and ref|NP_(—)244119.1|, ref|ZP_(—)02850022.1|,ref|YP_(—)815236.1|, ref|NP_(—)965472.1|, and ref|YP_(—)001422284.1|X(SEQ ID Nos: 2808-2812 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 157 depicts a sequence alignment between SEQ ID NO: 2823(RAAC01817) and ref|ZP_(—)02326197.1|, ref|YP_(—)001212789.1|,ref|ZP_(—)01666637.1|, ref|YP_(—)001211675.1|, andref|ZP_(—)01126596.1|X (SEQ ID Nos: 2825-2829 respectively). Amino acidsconserved among all sequences are indicated by a “*” and generallyconserved amino acids are indicated by a “:”.

FIG. 158 depicts a sequence alignment between SEQ ID NO: 2840(RAAC01840) and ref|YP_(—)001514193.1|, ref|YP_(—)517020.1|,ref|YP_(—)001317996.1|, ref|YP_(—)001090064.1|, andref|ZP_(—)01995293.1|X (SEQ ID Nos: 2842-2846 respectively). Amino acidsconserved among all sequences are indicated by a “*” and generallyconserved amino acids are indicated by a “:”.

FIG. 159 depicts a sequence alignment between SEQ ID NO: 2857(RAAC01875) and ref|YP_(—)644758.1|, ref|YP_(—)604970.1|,ref|YP_(—)076566.1|, ref|NP_(—)295807.1|, and ref|YP_(—)357266.1|X (SEQID Nos: 2859-2863 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 160 depicts a sequence alignment between SEQ ID NO: 2874(RAAC02539) and ref|ZP_(—)01034116.1|, gb|ABY83631.1|,ref|ZP_(—)02297879.1|, ref|YP_(—)208280.1|, and ref|YP_(—)001516905.1|X(SEQ ID Nos: 2876-2880 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 161 depicts a sequence alignment between SEQ ID NO: 2891(RAAC02543) and ref|ZP_(—)01042597.1|, gb|ABO14793.1|,ref|YP_(—)001443312.1|, ref|ZP_(—)01815366.1|, and ref|YP_(—)205672.1|X(SEQ ID Nos: 2893-2897 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 162 depicts a sequence alignment between SEQ ID NO: 2908(RAAC02564) and ref|ZP_(—)01631840.1|, gb|AAZ73681.1|,ref|YP_(—)001633718.1|, ref|YP_(—)001275109.1|, andref|ZP_(—)01514022.1|X (SEQ ID Nos: 2910-2914 respectively). Amino acidsconserved among all sequences are indicated by a “*” and generallyconserved amino acids are indicated by a “:”.

FIG. 163 depicts a sequence alignment between SEQ ID NO: 2925(RAAC02566) and ref|YP_(—)853610.1|, ref|YP_(—)512277.1|,ref|YP_(—)001038857.1|, ref|ZP_(—)01959153.1|, andref|YP_(—)001662865.1|X (SEQ ID Nos: 2927-2931 respectively). Aminoacids conserved among all sequences are indicated by a “*” and generallyconserved amino acids are indicated by a “:”.

FIG. 164 depicts a sequence alignment between SEQ ID NO: 2942(RAAC02589) and ref|ZP_(—)00591928.1|, ref|YP_(—)001003150.1|,ref|NP_(—)046614.1|, ref|YP_(—)375842.1|, and ref|YP_(—)001131112.1|X(SEQ ID Nos: 2944-2948 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 165 depicts a sequence alignment between SEQ ID NO: 2959(RAAC02045) and ref|ZP_(—)02848186.1|, ref|YP_(—)080797.1|,ref|YP_(—)001127101.1|, ref|YP_(—)148915.1|, and ref|YP_(—)001488316.1|X(SEQ ID Nos: 2961-2965 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 166 depicts a sequence alignment between SEQ ID NO: 2976(RAAC02635) and ref|YP_(—)866618.1|, ref|YP_(—)342704.1|,ref|NP_(—)384606.1|, ref|YP_(—)115396.1|, and ref|YP_(—)911114.1|X (SEQID Nos: 2978-2982 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 167 depicts a sequence alignment between SEQ ID NO: 2993(RAAC00088) and ref|NP_(—)244431.1|, ref|ZP_(—)02848186.1|,ref|ZP_(—)02171648.1|, ref|ZP_(—)02210735.1|, and ref|YP_(—)176521.1|X(SEQ ID Nos: 2995-2999 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 168 depicts a sequence alignment between SEQ ID NO: 3010(RAAC00111) and ref|YP_(—)146448.1|, ref|NP_(—)977168.1|,ref|ZP_(—)02261191.1|, ref|ZP_(—)02848045.1|, and ref|NP_(—)981890.1|X(SEQ ID Nos: 3012-3016 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 169 depicts a sequence alignment between SEQ ID NO: 3027(RAAC03161) and ref|ZP_(—)01898092.1|, ref|ZP_(—)01221581.1|,ref|YP_(—)128524.1|, ref|ZP_(—)01236658.1|, and ref|ZP_(—)01161642.1|X(SEQ ID Nos: 3029-3033 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 170 depicts a sequence alignment between SEQ ID NO: 3044(RAAC03110) and ref|YP_(—)001567845.1|, gb|ACA00199.1|,ref|ZP_(—)01731959.1|, ref|NP_(—)490383.1|, and ref|YP_(—)319960.1|X(SEQ ID Nos: 3046-3050 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 171 depicts a sequence alignment between SEQ ID NO: 3061(RAAC03810) and emb|CAP00374.1|, ref|YP_(—)001038857.1|,ref|NP_(—)852746.1|, ref|ZP_(—)01959153.1|, and ref|YP_(—)853610.1|X(SEQ ID Nos: 3063-3067 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 172 depicts a sequence alignment between SEQ ID NO: 3078(RAAC03316) and emb|CAP00374.1|, ref|YP_(—)001038857.1|,ref|ZP_(—)02327844.1|, ref|NP_(—)852746.1|, and ref|ZP_(—)01959153.1|X(SEQ ID Nos: 3080-3084 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 173 depicts a sequence alignment between SEQ ID NO: 3095(RAAC03018) and ref|YP_(—)148854.1|, ref|YP_(—)001127043.1|,ref|YP_(—)080555.1|, ref|NP_(—)693306.1|, and ref|ZP_(—)02170973.1|X(SEQ ID Nos: 3097-3101 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 174 depicts a sequence alignment between SEQ ID NO: 3112(RAAC02718) and ref|YP_(—)001178926.1|, ref|YP_(—)001180877.1|,ref|YP_(—)001219837.1|, ref|YP_(—)001211485.1|, andref|YP_(—)001211518.1|X (SEQ ID Nos: 3114-3118 respectively). Aminoacids conserved among all sequences are indicated by a “*” and generallyconserved amino acids are indicated by a “:”.

FIG. 175 depicts a sequence alignment between SEQ ID NO: 3129(RAAC01115) and ref|YP_(—)001035109.1|, ref|ZP_(—)01819917.1|,ref|YP_(—)001450442.1|, ref|ZP_(—)01818216.1|, andref|ZP_(—)01830009.1|X (SEQ ID Nos: 3131-3135 respectively). Amino acidsconserved among all sequences are indicated by a “*” and generallyconserved amino acids are indicated by a “:”.

FIG. 176 depicts a sequence alignment between SEQ ID NO: 3146(RAAC01119) and ref|NP_(—)622644.1|, ref|YP_(—)001664955.1|,ref|YP_(—)076287.1|, ref|ZP_(—)02616274.1|, and ref|YP_(—)001255422.1|X(SEQ ID Nos: 3148-3152 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 177 depicts a sequence alignment between SEQ ID NO: 3163(RAAC00203) and ref|YP_(—)001433837.1|, ref|YP_(—)001276310.1|,ref|ZP_(—)01514627.1|, ref|YP_(—)001636906.1|, andref|YP_(—)001102862.1|X (SEQ ID Nos: 3165-3169 respectively). Aminoacids conserved among all sequences are indicated by a “*” and generallyconserved amino acids are indicated by a “:”.

FIG. 178 depicts a sequence alignment between SEQ ID NO: 3180(RAAC01413) and ref|ZP_(—)02849297.1|, ref|NP_(—)815033.1|,ref|YP_(—)796463.1|, ref|NP_(—)862606.1|, and ref|ZP_(—)00604769.1|X(SEQ ID Nos: 3182-3186 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 179 depicts a sequence alignment between SEQ ID NO: 3197(RAAC01435) and ref|YP_(—)359065.1|, ref|YP_(—)145897.2|,ref|NP_(—)387932.1|, ref|YP_(—)001124176.1|, and sp|P42816|KPRS_BACCLX(SEQ ID Nos: 3199-3203 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 180 depicts a sequence alignment between SEQ ID NO: 3214(RAAC01442) and ref|ZP_(—)02170919.1|, ref|YP_(—)535778.1|,ref|ZP_(—)01862118.1|, ref|NP_(—)692713.1|, and ref|YP_(—)359077.1|X(SEQ ID Nos: 3216-3220 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 181 depicts a sequence alignment between SEQ ID NO: 3231(RAAC00895) and ref|NP_(—)846740.1|, ref|ZP_(—)00238564.1|,ref|ZP_(—)02583512.1|, ref|NP_(—)834002.1|, and ref|ZP_(—)02604064.1|X(SEQ ID Nos: 3233-3237 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

FIG. 182 depicts a sequence alignment between SEQ ID NO: 3248(RAAC03475) and ref|ZP_(—)02442523.1|, ref|YP_(—)001210714.1|,ref|YP_(—)001180871.1|, ref|YP_(—)001662345.1|, andref|YP_(—)001114120.1|X (SEQ ID Nos: 3250-3254 respectively). Aminoacids conserved among all sequences are indicated by a “*” and generallyconserved amino acids are indicated by a “:”.

FIG. 183 depicts a sequence alignment between SEQ ID NO: 3265(RAAC03560) and ref|YP_(—)001114120.1|, ref|NP_(—)242309.1|,ref|YP_(—)146346.1|, ref|YP_(—)146347.1|, and ref|YP_(—)001662345.1|X(SEQ ID Nos: 3267-3271 respectively). Amino acids conserved among allsequences are indicated by a “*” and generally conserved amino acids areindicated by a “:”.

DETAILED DESCRIPTION OF THE INVENTION

Embodiments of the invention include genes and associated proteinsrelated to genetic recombination from the thermoacidophileAlicyclobacillus acidocaldarius. Coding sequences for genes related torecombination were determined from sequence information generated fromsequencing the genome of Alicyclobacillus acidocaldarius. These genesand proteins may represent targets and/or elements of transformationsystems or vectors for genetic engineering for introducing nucleotidesequences of interest into Alicyclobacillus acidocaldarius, Grampositive thermophiles, or other organisms. Non-limiting examples ofnucleotide sequences found within the genome of Alicyclobacillusacidocaldarius, and amino acids coded thereby, associated withrecombination are listed in the sequence listing. Examples of thesenucleotide sequences and the proteins they encode can be found inTable 1. Proteins related to recombination may be, without limitation,one of any of the following: ATP-dependent DNA helicase recG,ATP-dependent DNA ligase, ATP-dependent endopeptidase clp proteolyticsubunit, Chromosome partitioning protein, Crossover junctionendodeoxyribonuclease ruvC, Deoxyuridine 5′-triphosphatenucleotidohydrolase, DNA adenine methylase, DNA helicase, DNAintegration/recombination/inversion, DNA polymerase I, DNA polymeraseIII beta chain, DNA polymerase IV, DNA primase, DNA repair protein radC,DNA replication and repair protein recF, DNA replication protein dnaD,DNA topoisomerase I, DNA/RNA helicase (DEAD/DEAH box family),DNA-binding protein HU, Fe—S oxidoreductase, Glycerophosphoryl diesterphosphodiesterase, HNH endonuclease family protein, Holliday junctionDNA helicase ruvB, Integrase/recombinase (XerC/CodV family),Ligase/carboxyalse family protein, LtrC-like protein, Macrolide-effluxprotein, NAD-dependent DNA ligase, Nicotinate phosphoribosyltransferase,nodulin-26, Phage antirepressor protein, Phage protein, PhosphinothricinN-acetyltransferase, Phosphohydrolase, RecA protein, Recombinationprotein recR, Replicative DNA helicase, Ribonucleoside-diphosphatereductase alpha chain, Ribonucleoside-diphosphate reductase beta chain,Ribose-phosphate pyrophosphokinase, Serine/threonine proteinphosphatase, Single-strand DNA binding protein, Single-strandedDNA-binding protein, Site-specific recombinase, Site-specificresolvase/integrase, Thioredoxin, Thymidine kinase, Transcriptionalregulator/Lex A repressor, Transcriptional regulator, Cro/CI family,Transposase, TRSE protein, Two-component response regulator, Type IIrestriction-modification system methylation subunit, and others.

Embodiments of the invention relate in part to the gene sequences and/orprotein sequences comprising genes and/or proteins of Alicyclobacillusacidocaldarius. Genes and proteins included are those which play a rolein genetic manipulation. Intracellular enzyme activities may bethermophilic and/or acidophilic in nature and general examples ofsimilar genes are described in the literature. Classes of genes,sequences, enzymes and factors include, but are not limited to, thoselisted in Table 1. FIGS. 1-183 provide sequence alignments betweenpolypeptide sequences of the present invention and closely relatedproteins. Areas of high homology between the polypeptide sequences ofthe present invention and closely related proteins are indicative offunctionality for polypeptide sequences of the present invention for thesame purposes as the sequences to which they are aligned.

TABLE 1 Alicyclobacillus acidocaldarius coding sequences related torecombination Reference Protein Sequence Coding Sequence FunctionRAAC02304 SEQ ID NO: 137 SEQ ID NO: 138 Two-component response regulatorRAAC02309 SEQ ID NO: 222 SEQ ID NO: 223 TRSE PROTEIN RAAC02315 SEQ IDNO: 324 SEQ ID NO: 325 Chromosome partitioning protein parA RAAC02319SEQ ID NO: 409 SEQ ID NO: 410 Transposase RAAC02321 SEQ ID NO: 443 SEQID NO: 444 Ligase/carboxyalse family protein RAAC02324 SEQ ID NO: 494SEQ ID NO: 495 Fe-S OXIDOREDUCTASE RAAC02326 SEQ ID NO: 511 SEQ ID NO:512 nodulin-26 RAAC02332 SEQ ID NO: 562 SEQ ID NO: 563 PhosphinothricinN-acetyltransferase RAAC02292 SEQ ID NO: 647 SEQ ID NO: 648Transcriptional regulator/Lex A repressor RAAC02340 SEQ ID NO: 715 SEQID NO: 716 LtrC-like protein RAAC02341 SEQ ID NO: 732 SEQ ID NO: 733 DNArepair protein radC RAAC02342 SEQ ID NO: 749 SEQ ID NO: 750 DNApolymerase III, beta chain RAAC02348 SEQ ID NO: 834 SEQ ID NO: 835Single-stranded DNA-binding protein RAAC02358 SEQ ID NO: 1004 SEQ ID NO:1005 DNA/RNA helicase (DEAD/DEAH box family) RAAC02361 SEQ ID NO: 1038SEQ ID NO: 1039 DNA helicase II RAAC02363 SEQ ID NO: 1072 SEQ ID NO:1073 DNA polymerase IV RAAC02318 SEQ ID NO: 1327 SEQ ID NO: 1328Transposase RAAC02319 SEQ ID NO: 1344 SEQ ID NO: 1345 TransposaseRAAC02333 SEQ ID NO: 1361 SEQ ID NO: 1362 DNAintegration/recombination/inversion protein RAAC03173 SEQ ID NO: 1429SEQ ID NO: 1430 Transposase RAAC00757 SEQ ID NO: 1463 SEQ ID NO: 1464Holliday junction DNA helicase ruvB RAAC00756 SEQ ID NO: 1480 SEQ ID NO:1481 Holliday junction DNA helicase ruvB RAAC00755 SEQ ID NO: 1497 SEQID NO: 1498 Crossover junction endodeoxyribonuclease ruvC RAAC01468 SEQID NO: 1514 SEQ ID NO: 1515 Site-specific recombinase RAAC03178 SEQ IDNO: 1531 SEQ ID NO: 1532 Transposase RAAC01937 SEQ ID NO: 1548 SEQ IDNO: 1549 DNA polymerase I RAAC01372 SEQ ID NO: 1565 SEQ ID NO: 1566Transposase RAAC02377 SEQ ID NO: 1599 SEQ ID NO: 1600 TransposaseRAAC00037 SEQ ID NO: 1650 SEQ ID NO: 1651 ATP-dependent DNA helicaserecG RAAC00054 SEQ ID NO: 1667 SEQ ID NO: 1668 DNA replication andrepair protein recF RAAC03102 SEQ ID NO: 1684 SEQ ID NO: 1685Transposase RAAC03103 SEQ ID NO: 1701 SEQ ID NO: 1702 TransposaseRAAC03341 SEQ ID NO: 1718 SEQ ID NO: 1719 Transposase RAAC02852 SEQ IDNO: 1786 SEQ ID NO: 1787 DNA topoisomerase I (EC 1280.1374.1276.1277)RAAC02854 SEQ ID NO: 1803 SEQ ID NO: 1804 Integrase/recombinase(XerC/CodV family) RAAC03166 SEQ ID NO: 1820 SEQ ID NO: 1821 TransposaseRAAC02961 SEQ ID NO: 1854 SEQ ID NO: 1855 DNAintegration/recombination/inversion protein RAAC02202 SEQ ID NO: 1871SEQ ID NO: 1872 NAD-dependent DNA ligase RAAC03682 SEQ ID NO: 1888 SEQID NO: 1889 Transposase RAAC02738 SEQ ID NO: 1939 SEQ ID NO: 1940 DNAintegration/recombination/inversion protein RAAC02514 SEQ ID NO: 1956SEQ ID NO: 1957 Transposase RAAC02515 SEQ ID NO: 1973 SEQ ID NO: 1974Transposase RAAC02530 SEQ ID NO: 1990 SEQ ID NO: 1991 TransposaseRAAC02533 SEQ ID NO: 2007 SEQ ID NO: 2008 Transposase RAAC02534 SEQ IDNO: 2024 SEQ ID NO: 2025 Transposase RAAC02562 SEQ ID NO: 2041 SEQ IDNO: 2042 Transposase RAAC03229 SEQ ID NO: 2058 SEQ ID NO: 2059Transposase RAAC00160 SEQ ID NO: 2092 SEQ ID NO: 2093 ATP-dependent DNAhelicase recG RAAC03182 SEQ ID NO: 2109 SEQ ID NO: 2110 TransposaseRAAC03163 SEQ ID NO: 2126 SEQ ID NO: 2127 Transposase RAAC01387 SEQ IDNO: 2143 SEQ ID NO: 2144 Transposase RAAC01171 SEQ ID NO: 2279 SEQ IDNO: 2280 Transposase RAAC03825 SEQ ID NO: 2296 SEQ ID NO: 2297Transposase RAAC03826 SEQ ID NO: 2313 SEQ ID NO: 2314 TransposaseRAAC02717 SEQ ID NO: 2330 SEQ ID NO: 2331 Transposase RAAC01155 SEQ IDNO: 2347 SEQ ID NO: 2348 Recombination protein recR RAAC03145 SEQ ID NO:2364 SEQ ID NO: 2365 Transposase RAAC03325 SEQ ID NO: 2381 SEQ ID NO:2382 Transposase RAAC03376 SEQ ID NO: 2398 SEQ ID NO: 2399 TransposaseRAAC02657 SEQ ID NO: 2415 SEQ ID NO: 2416 Type IIrestriction-modification system methylation subunit RAAC01373 SEQ ID NO:2432 SEQ ID NO: 2433 Site-specific resolvase/integrase RAAC00337 SEQ IDNO: 2449 SEQ ID NO: 2450 DNA adenine methylase RAAC00506 SEQ ID NO: 2466SEQ ID NO: 2467 Serine/threonine protein phosphatase RAAC00022 SEQ IDNO: 2483 SEQ ID NO: 2484 Replicative DNA helicase RAAC00027 SEQ ID NO:2500 SEQ ID NO: 2501 Single-strand DNA binding protein RAAC01051 SEQ IDNO: 2517 SEQ ID NO: 2518 Ribonucleoside-diphosphate reductase alphachain RAAC01050 SEQ ID NO: 2534 SEQ ID NO: 2535Ribonucleoside-diphosphate reductase beta chain RAAC01009 SEQ ID NO:2551 SEQ ID NO: 2552 DNA replication protein dnaD RAAC00998 SEQ ID NO:2568 SEQ ID NO: 2569 ATP-dependent DNA ligase RAAC02359 SEQ ID NO: 2585SEQ ID NO: 2586 DNA-binding protein HU RAAC00997 SEQ ID NO: 2602 SEQ IDNO: 2603 ATP-dependent DNA ligase RAAC02419 SEQ ID NO: 2619 SEQ ID NO:2620 Site-specific recombinase RAAC02417 SEQ ID NO: 2636 SEQ ID NO: 2637Transcriptional regulator, Cro/CI family RAAC03180 SEQ ID NO: 2653 SEQID NO: 2654 Transcriptional regulator, Cro/CI family RAAC03224 SEQ IDNO: 2687 SEQ ID NO: 2688 Phage antirepressor protein RAAC02915 SEQ IDNO: 2704 SEQ ID NO: 2705 Deoxyuridine 5′-triphosphatenucleotidohydrolase RAAC02943 SEQ ID NO: 2721 SEQ ID NO: 2722 RecAprotein RAAC02234 SEQ ID NO: 2738 SEQ ID NO: 2739 Serine/threonineprotein phosphatase RAAC01662 SEQ ID NO: 2755 SEQ ID NO: 2756Macrolide-efflux protein RAAC02171 SEQ ID NO: 2772 SEQ ID NO: 2773Phosphohydrolase RAAC01696 SEQ ID NO: 2789 SEQ ID NO: 2790 ThioredoxinRAAC01724 SEQ ID NO: 2806 SEQ ID NO: 2807 Thioredoxin RAAC01817 SEQ IDNO: 2823 SEQ ID NO: 2824 Site-specific recombinase RAAC01840 SEQ ID NO:2840 SEQ ID NO: 2841 Nicotinate phosphoribosyltransferase RAAC01875 SEQID NO: 2857 SEQ ID NO: 2858 Glycerophosphoryl diester phosphodiesteraseRAAC02539 SEQ ID NO: 2874 SEQ ID NO: 2875 Type IIrestriction-modification system methylation subunit RAAC02543 SEQ ID NO:2891 SEQ ID NO: 2892 DNA adenine methylase RAAC02564 SEQ ID NO: 2908 SEQID NO: 2909 DNA adenine methylase RAAC02566 SEQ ID NO: 2925 SEQ ID NO:2926 Phage antirepressor protein RAAC02589 SEQ ID NO: 2942 SEQ ID NO:2943 DNA-binding protein HU RAAC02045 SEQ ID NO: 2959 SEQ ID NO: 2960ATP-dependent endopeptidase clp proteolytic subunit clpP RAAC02635 SEQID NO: 2976 SEQ ID NO: 2977 Type II restriction-modification systemmethylation subunit RAAC00088 SEQ ID NO: 2993 SEQ ID NO: 2994ATP-dependent endopeptidase clp proteolytic subunit clpP RAAC00111 SEQID NO: 3010 SEQ ID NO: 3011 Site-specific recombinase RAAC03161 SEQ IDNO: 3027 SEQ ID NO: 3028 DNA adenine methylase RAAC03110 SEQ ID NO: 3044SEQ ID NO: 3045 HNH endonuclease family protein RAAC03316 SEQ ID NO:3078 SEQ ID NO: 3079 Phage antirepressor protein RAAC03018 SEQ ID NO:3095 SEQ ID NO: 3096 Thioredoxin RAAC02718 SEQ ID NO: 3112 SEQ ID NO:3113 Phage protein RAAC01115 SEQ ID NO: 3129 SEQ ID NO: 3130 Thymidinekinase RAAC01119 SEQ ID NO: 3146 SEQ ID NO: 3147 Glycerophosphoryldiester phosphodiesterase RAAC00203 SEQ ID NO: 3163 SEQ ID NO: 3164Ribonucleoside-diphosphate reductase beta chain RAAC01413 SEQ ID NO:3180 SEQ ID NO: 3181 Serine/threonine protein phosphatase RAAC01435 SEQID NO: 3197 SEQ ID NO: 3198 Ribose-phosphate pyrophosphokinase RAAC01442SEQ ID NO: 3214 SEQ ID NO: 3215 DNA-binding protein HU RAAC00895 SEQ IDNO: 3231 SEQ ID NO: 3232 DNA primase RAAC03475 SEQ ID NO: 3248 SEQ IDNO: 3249 Transcriptional regulator, Cro/CI family

The present invention relates to nucleotides sequences comprisingisolated and/or purified nucleotide sequences of the genome ofAlicyclobacillus acidocaldarius selected from the sequences of SEQ IDNos. 2, 19, 36, 53, 70, 87, 104, 121, 138, 155, 172, 189, 206, 223, 240,257, 274, 291, 308, 325, 359, 376, 410, 427, 444, 512, 529, 546, 597,614, 648, 665, 682, 699, 716, 733, 750, 767, 784, 818, 835, 852, 886,903, 920, 937, 954, 971, 988, 1005, 1039, 1056, 1073, 1090, 1107, 1124,1141, 1158, 1175, 1192, 1209, 1226, 1243, 1260, 1277, 1294, 1311, 1328,1345, 1362, 1379, 1396, 1413, 1430, 1447, 1464, 1481, 1498, 1515, 1532,1549, 1566, 1583, 1600, 1634, 1651, 1685, 1702, 1719, 1787, 1804, 1821,1872, 1889, 1923, 1940, 1957, 1974, 1991, 2008, 2025, 2042, 2059, 2093,2110, 2127, 2161, 2178, 2195, 2212, 2229, 2246, 2263, 2280, 2314, 2331,2348, 2365, 2382, 2399, 2416, 2433, 2450, 2467, 2484, 2501, 2518, 2535,2552, 2569, 2586, 2603, 2620, 2637, 2654, 2671, 2688, 2705, 2722, 2739,2773, 2790, 2824, 2841, 2858, 2926, 2943, 2960, 3011, 3028, 3062, 3079,3096, 3113, 3130, 3147, 3164, 3181, 3198, 3232, 3249, 3266, and 3283 orone of their fragments.

The present invention likewise relates to isolated and/or purifiednucleotide sequences, characterized in that they comprise at least oneof: a) a nucleotide sequence of at least one of the sequences of SEQ IDNos. 2, 19, 36, 53, 70, 87, 104, 121, 138, 155, 172, 189, 206, 223, 240,257, 274, 291, 308, 325, 359, 376, 410, 427, 444, 512, 529, 546, 597,614, 648, 665, 682, 699, 716, 733, 750, 767, 784, 818, 835, 852, 886,903, 920, 937, 954, 971, 988, 1005, 1039, 1056, 1073, 1090, 1107, 1124,1141, 1158, 1175, 1192, 1209, 1226, 1243, 1260, 1277, 1294, 1311, 1328,1345, 1362, 1379, 1396, 1413, 1430, 1447, 1464, 1481, 1498, 1515, 1532,1549, 1566, 1583, 1600, 1634, 1651, 1685, 1702, 1719, 1787, 1804, 1821,1872, 1889, 1923, 1940, 1957, 1974, 1991, 2008, 2025, 2042, 2059, 2093,2110, 2127, 2161, 2178, 2195, 2212, 2229, 2246, 2263, 2280, 2314, 2331,2348, 2365, 2382, 2399, 2416, 2433, 2450, 2467, 2484, 2501, 2518, 2535,2552, 2569, 2586, 2603, 2620, 2637, 2654, 2671, 2688, 2705, 2722, 2739,2773, 2790, 2824, 2841, 2858, 2926, 2943, 2960, 3011, 3028, 3062, 3079,3096, 3113, 3130, 3147, 3164, 3181, 3198, 3232, 3249, 3266, and 3283 orone of their fragments; b) a nucleotide sequence homologous to anucleotide sequence such as defined in a); c) a nucleotide sequencecomplementary to a nucleotide sequence such as defined in a) or b), anda nucleotide sequence of their corresponding RNA; d) a nucleotidesequence capable of hybridizing under stringent conditions with asequence such as defined in a), b) or c); e) a nucleotide sequencecomprising a sequence such as defined in a), b), c) or d); and f) anucleotide sequence modified by a nucleotide sequence such as defined ina), b), c), d) or e).

Nucleotide, polynucleotide, or nucleic acid sequence will be understoodaccording to the present invention as meaning both a double-stranded orsingle-stranded DNA in the monomeric and dimeric (so-called in tandem)forms and the transcription products of said DNAs.

Aspects of the invention relate to nucleotide sequences which it hasbeen possible to isolate, purify or partially purify, starting fromseparation methods such as, for example, ion-exchange chromatography, byexclusion based on molecular size, or by affinity, or alternativelyfractionation techniques based on solubility in different solvents, orstarting from methods of genetic engineering such as amplification,cloning, and subcloning, it being possible for the sequences of theinvention to be carried by vectors.

Isolated and/or purified nucleotide sequence fragment according to theinvention will be understood as designating any nucleotide fragment ofthe genome of Alicyclobacillus acidocaldarius, and may include, by wayof non-limiting examples, length of at least 8, 12, 20 25, 50, 75, 100,200, 300, 400, 500, 1000, or more, consecutive nucleotides of thesequence from which it originates.

Specific fragment of an isolated and/or purified nucleotide sequenceaccording to the invention will be understood as designating anynucleotide fragment of the genome of Alicyclobacillus acidocaldarius,having, after alignment and comparison with the corresponding fragmentsof genomic sequences of Alicyclobacillus acidocaldarius, at least onenucleotide or base of different nature.

Homologous isolated and/or purified nucleotide sequence in the sense ofthe present invention is understood as meaning isolated and/or purifieda nucleotide sequence having at least a percentage identity with thebases of a nucleotide sequence according to the invention of at leastabout 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%,93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.6%, or 99.7%, thispercentage being purely statistical and it being possible to distributethe differences between the two nucleotide sequences at random and overthe whole of their length.

Specific homologous nucleotide sequence in the sense of the presentinvention is understood as meaning a homologous nucleotide sequencehaving at least one nucleotide sequence of a specific fragment, such asdefined above. Said “specific” homologous sequences can comprise, forexample, the sequences corresponding to the genomic sequence or to thesequences of its fragments representative of variants of the genome ofAlicyclobacillus acidocaldarius. These specific homologous sequences canthus correspond to variations linked to mutations within strains ofAlicyclobacillus acidocaldarius, and especially correspond totruncations, substitutions, deletions and/or additions of at least onenucleotide. Said homologous sequences can likewise correspond tovariations linked to the degeneracy of the genetic code.

The term “degree or percentage of sequence homology” refers to “degreeor percentage of sequence identity between two sequences after optimalalignment” as defined in the present application.

Two amino-acids or nucleotide sequences are said to be “identical” ifthe sequence of amino-acids or nucleotide residues, in the two sequencesis the same when aligned for maximum correspondence as described below.Sequence comparisons between two (or more) peptides or polynucleotidesare typically performed by comparing sequences of two optimally alignedsequences over a segment or “comparison window” to identify and comparelocal regions of sequence similarity. Optimal alignment of sequences forcomparison may be conducted by the local homology algorithm of Smith andWaterman, J Mol Biol, 147, 195-197, by the homology alignment algorithmof Needleman and Wunsch, J. Mol. Biol. 48: 443 (1970), by the search forsimilarity method of Pearson and Lipman, Proc. Natl. Acad. Sci. (U.S.A.)85: 2444 (1988), by computerized implementation of these algorithms(GAP, BESTFIT, FASTA, and TFASTA in the Wisconsin Genetics SoftwarePackage, Genetics Computer Group (GCG), 575 Science Dr., Madison, Wis.),or by visual inspection.

“Percentage of sequence identity” (or degree of identity) is determinedby comparing two optimally aligned sequences over a comparison window,where the portion of the peptide or polynucleotide sequence in thecomparison window may comprise additions or deletions (i.e., gaps) ascompared to the reference sequence (which does not comprise additions ordeletions) for optimal alignment of the two sequences. The percentage iscalculated by determining the number of positions at which the identicalamino-acid residue or nucleic acid base occurs in both sequences toyield the number of matched positions, dividing the number of matchedpositions by the total number of positions in the window of comparisonand multiplying the result by 100 to yield the percentage of sequenceidentity.

The definition of sequence identity given above is the definition thatwould be used by one of skill in the art. The definition by itself doesnot need the help of any algorithm, said algorithms being helpful onlyto achieve the optimal alignments of sequences, rather than thecalculation of sequence identity.

From the definition given above, it follows that there is a well definedand only one value for the sequence identity between two comparedsequences which value corresponds to the value obtained for the best oroptimal alignment.

In the BLAST N or BLAST P “BLAST 2 sequence”, software which isavailable in the web siteworldwideweb.blast.ncbi.nlm.nih.gov/Blast.cgi?PAGE_TYPE=BlastSearch&PROG_DEF=blastn&BLAST_PROG_DEF=megaBlast&SHOW_DEFAULTS=on&BLAST_SPEC=blast2seq&LINK_LOC=align2seq

, and habitually used by the inventors and in general by the skilledperson for comparing and determining the identity between two sequences,gap cost which depends on the sequence length to be compared is directlyselected by the software (i.e. 11.2 for substitution matrix BLOSUM-62for length>85).

Complementary nucleotide sequence of a sequence of the invention isunderstood as meaning any DNA whose nucleotides are complementary tothose of the sequence of the invention, and whose orientation isreversed (antisense sequence).

Hybridization under conditions of stringency with a nucleotide sequenceaccording to the invention is understood as meaning hybridization underconditions of temperature and ionic strength chosen in such a way thatthey allow the maintenance of the hybridization between two fragments ofcomplementary DNA.

By way of illustration, conditions of great stringency of thehybridization step with the aim of defining the nucleotide fragmentsdescribed above are advantageously the following.

The hybridization is carried out at a preferential temperature of 65° C.in the presence of SSC buffer, 1×SSC corresponding to 0.15 M NaCl and0.05 M Na citrate. The washing steps, for example, can be the following:2×SSC, at ambient temperature followed by two washes with 2×SSC, 0.5%SDS at 65° C.; 2×0.5×SSC, 0.5% SDS; at 65° C. for 10 minutes each.

The conditions of intermediate stringency, using, for example, atemperature of 42° C. in the presence of a 2×SSC buffer, or of lessstringency, for example a temperature of 37° C. in the presence of a2×SSC buffer, respectively require a globally less significantcomplementarity for the hybridization between the two sequences.

The stringent hybridization conditions described above for apolynucleotide with a size of approximately 350 bases will be adapted bythe person skilled in the art for oligonucleotides of greater or smallersize, according to the teaching of Sambrook et al., 1989.

Among the isolated and/or purified nucleotide sequences according to theinvention, are those which can be used as a primer or probe in methodsallowing the homologous sequences according to the invention to beobtained, these methods, such as the polymerase chain reaction (PCR),nucleic acid cloning, and sequencing, being well known to the personskilled in the art.

The terms “recombination,” “genetic recombination,” and “geneticengineering” are used interchangeable herein and refer to the process bywhich a first stretch of one or more nucleic acids are removed from oradded to a second stretch of nucleic acids. Such addition or removal mayoccur in vivo or ex vivo. The terms also refer to the introduction ofcoding sequences or genes into cells or organisms for the purposes ofexpression or regulation of other nucleotide sequences or polypeptides.The terms further refer to the alteration of nucleic acid structure ortopology. By way of non-limiting example, altering the “twist,”supercoiling, helicity, separation and/or annealing of a nucleotidesequence.

Among the isolated and/or purified nucleotide sequences according to theinvention, those are again preferred which can be used as a primer orprobe in methods allowing the presence of SEQ ID Nos. 2, 19, 36, 53, 70,87, 104, 121, 138, 155, 172, 189, 206, 223, 240, 257, 274, 291, 308,325, 359, 376, 410, 427, 444, 512, 529, 546, 597, 614, 648, 665, 682,699, 716, 733, 750, 767, 784, 818, 835, 852, 886, 903, 920, 937, 954,971, 988, 1005, 1039, 1056, 1073, 1090, 1107, 1124, 1141, 1158, 1175,1192, 1209, 1226, 1243, 1260, 1277, 1294, 1311, 1328, 1345, 1362, 1379,1396, 1413, 1430, 1447, 1464, 1481, 1498, 1515, 1532, 1549, 1566, 1583,1600, 1634, 1651, 1685, 1702, 1719, 1787, 1804, 1821, 1872, 1889, 1923,1940, 1957, 1974, 1991, 2008, 2025, 2042, 2059, 2093, 2110, 2127, 2161,2178, 2195, 2212, 2229, 2246, 2263, 2280, 2314, 2331, 2348, 2365, 2382,2399, 2416, 2433, 2450, 2467, 2484, 2501, 2518, 2535, 2552, 2569, 2586,2603, 2620, 2637, 2654, 2671, 2688, 2705, 2722, 2739, 2773, 2790, 2824,2841, 2858, 2926, 2943, 2960, 3011, 3028, 3062, 3079, 3096, 3113, 3130,3147, 3164, 3181, 3198, 3232, 3249, 3266, and 3283, and one of theirfragments, or one of their variants such as defined below to bediagnosed.

The nucleotide sequence fragments according to the invention can beobtained, for example, by specific amplification, such as PCR, or afterdigestion with appropriate restriction enzymes of nucleotide sequencesaccording to the invention, these methods in particular being describedin the work of Sambrook et al., 1989. Such representative fragments canlikewise be obtained by chemical synthesis according to methods wellknown to persons of ordinary skill in the art.

Modified nucleotide sequence will be understood as meaning anynucleotide sequence obtained by mutagenesis according to techniques wellknown to the person skilled in the art, and containing modificationswith respect to the normal sequences according to the invention, forexample mutations in the regulatory and/or promoter sequences ofpolypeptide expression, especially leading to a modification of the rateof expression of said polypeptide or to a modulation of the replicativecycle.

Modified nucleotide sequence will likewise be understood as meaning anynucleotide sequence coding for a modified polypeptide such as definedbelow.

The present invention relates to nucleotide sequence comprising isolatedand/or purified nucleotide sequences of Alicyclobacillus acidocaldarius,characterized in that they are selected from the sequences SEQ ID Nos.2, 19, 36, 53, 70, 87, 104, 121, 138, 155, 172, 189, 206, 223, 240, 257,274, 291, 308, 325, 359, 376, 410, 427, 444, 512, 529, 546, 597, 614,648, 665, 682, 699, 716, 733, 750, 767, 784, 818, 835, 852, 886, 903,920, 937, 954, 971, 988, 1005, 1039, 1056, 1073, 1090, 1107, 1124, 1141,1158, 1175, 1192, 1209, 1226, 1243, 1260, 1277, 1294, 1311, 1328, 1345,1362, 1379, 1396, 1413, 1430, 1447, 1464, 1481, 1498, 1515, 1532, 1549,1566, 1583, 1600, 1634, 1651, 1685, 1702, 1719, 1787, 1804, 1821, 1872,1889, 1923, 1940, 1957, 1974, 1991, 2008, 2025, 2042, 2059, 2093, 2110,2127, 2161, 2178, 2195, 2212, 2229, 2246, 2263, 2280, 2314, 2331, 2348,2365, 2382, 2399, 2416, 2433, 2450, 2467, 2484, 2501, 2518, 2535, 2552,2569, 2586, 2603, 2620, 2637, 2654, 2671, 2688, 2705, 2722, 2739, 2773,2790, 2824, 2841, 2858, 2926, 2943, 2960, 3011, 3028, 3062, 3079, 3096,3113, 3130, 3147, 3164, 3181, 3198, 3232, 3249, 3266, and 3283 or one oftheir fragments.

Embodiments of the invention likewise relate to isolated and/or purifiednucleotide sequences characterized in that they comprise a nucleotidesequence selected from: a) at least one of a nucleotide sequence of SEQID Nos. 2, 19, 36, 53, 70, 87, 104, 121, 138, 155, 172, 189, 206, 223,240, 257, 274, 291, 308, 325, 359, 376, 410, 427, 444, 512, 529, 546,597, 614, 648, 665, 682, 699, 716, 733, 750, 767, 784, 818, 835, 852,886, 903, 920, 937, 954, 971, 988, 1005, 1039, 1056, 1073, 1090, 1107,1124, 1141, 1158, 1175, 1192, 1209, 1226, 1243, 1260, 1277, 1294, 1311,1328, 1345, 1362, 1379, 1396, 1413, 1430, 1447, 1464, 1481, 1498, 1515,1532, 1549, 1566, 1583, 1600, 1634, 1651, 1685, 1702, 1719, 1787, 1804,1821, 1872, 1889, 1923, 1940, 1957, 1974, 1991, 2008, 2025, 2042, 2059,2093, 2110, 2127, 2161, 2178, 2195, 2212, 2229, 2246, 2263, 2280, 2314,2331, 2348, 2365, 2382, 2399, 2416, 2433, 2450, 2467, 2484, 2501, 2518,2535, 2552, 2569, 2586, 2603, 2620, 2637, 2654, 2671, 2688, 2705, 2722,2739, 2773, 2790, 2824, 2841, 2858, 2926, 2943, 2960, 3011, 3028, 3062,3079, 3096, 3113, 3130, 3147, 3164, 3181, 3198, 3232, 3249, 3266, and3283 or one of their fragments or one of their fragments; b) anucleotide sequence of a specific fragment of a sequence such as definedin a); c) a homologous nucleotide sequence having at least 80% identitywith a sequence such as defined in a) or b); d) a complementarynucleotide sequence or sequence of RNA corresponding to a sequence suchas defined in a), b) or c); and e) a nucleotide sequence modified by asequence such as defined in a), b), c) or d).

Among the isolated and/or purified nucleotide sequences according to theinvention are the nucleotide sequences of SEQ ID Nos. 13-17, 30-34,47-51, 64-68, 81-85, 98-102, 115-119, 132-136, 149-153, 166-170,183-187, 200-204, 217-221, 234-238, 251-255, 268-272, 285-289, 302-306,319-323, 336-340, 353-357, 370-374, 387-391, 404-408, 421-425, 438-442,455-459, 472-476, 489-493, 506-510, 523-527, 540-544, 557-561, 574-578,591-595, 608-612, 625-629, 642-646, 659-663, 676-680, 693-697, 710-714,727-731, 744-748, 761-765, 778-782, 795-799, 812-816, 829-833, 846-850,863-867, 880-884, 897-901, 914-918, 931-935, 948-952, 965-969, 982-986,999-1003, 1016-1020, 1033-1037, 1050-1054, 1067-1071, 1084-1088,1101-1105, 1118-1122, 1135-1139, 1152-1156, 1169-1173, 1186-1190,1203-1207, 1220-1224, 1237-1241, 1254-1258, 1271-1275, 1288-1292,1305-1309, 1322-1326, 1339-1343, 1356-1360, 1373-1377, 1390-1394,1407-1411, 1424-1428, 1441-1445, 1458-1462, 1475-1479, 1492-1496,1509-1513, 1526-1530, 1543-1547, 1560-1564, 1577-1581, 1594-1598,1611-1615, 1628-1632, 1645-1649, 1662-1666, 1679-1683, 1696-1700,1713-1717, 1730-1734, 1747-1751, 1764-1768, 1781-1785, 1798-1802,1815-1819, 1832-1836, 1849-1853, 1866-1870, 1883-1887, 1900-1904,1917-1921, 1934-1938, 1951-1955, 1968-1972, 1985-1989, 2002-2006,2019-2023, 2036-2040, 2053-2057, 2070-2074, 2087-2091, 2104-2108,2121-2125, 2138-2142, 2155-2159, 2172-2176, 2189-2193, 2206-2210,2223-2227, 2240-2244, 2257-2261, 2274-2278, 2291-2295, 2308-2312,2325-2329, 2342-2346, 2359-2363, 2376-2380, 2393-2397, 2410-2414,2427-2431, 2444-2448, 2461-2465, 2478-2482, 2495-2499, 2512-2516,2529-2533, 2546-2550, 2563-2567, 2580-2584, 2597-2601, 2614-2618,2631-2635, 2648-2652, 2665-2669, 2682-2686, 2699-2703, 2716-2720,2733-2737, 2750-2754, 2767-2771, 2784-2788, 2801-2805, 2818-2822,2835-2839, 2852-2856, 2869-2873, 2886-2890, 2903-2907, 2920-2924,2937-2941, 2954-2958, 2971-2975, 2988-2992, 3005-3009, 3022-3026,3039-3043, 3056-3060, 3073-3077, 3090-3094, 3107-3111, 3124-3128,3141-3145, 3158-3162, 3175-3179, 3192-3196, 3209-3213, 3226-3230,3243-3247, 3260-3264, 3277-3281, and 3294-3298; or fragments thereof andany isolated and/or purified nucleotide sequences which have a homologyof at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%,92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.6%, or 99.7% identitywith the at least one of the sequences of SEQ ID Nos. 2, 19, 36, 53, 70,87, 104, 121, 138, 155, 172, 189, 206, 223, 240, 257, 274, 291, 308,325, 359, 376, 410, 427, 444, 512, 529, 546, 597, 614, 648, 665, 682,699, 716, 733, 750, 767, 784, 818, 835, 852, 886, 903, 920, 937, 954,971, 988, 1005, 1039, 1056, 1073, 1090, 1107, 1124, 1141, 1158, 1175,1192, 1209, 1226, 1243, 1260, 1277, 1294, 1311, 1328, 1345, 1362, 1379,1396, 1413, 1430, 1447, 1464, 1481, 1498, 1515, 1532, 1549, 1566, 1583,1600, 1634, 1651, 1685, 1702, 1719, 1787, 1804, 1821, 1872, 1889, 1923,1940, 1957, 1974, 1991, 2008, 2025, 2042, 2059, 2093, 2110, 2127, 2161,2178, 2195, 2212, 2229, 2246, 2263, 2280, 2314, 2331, 2348, 2365, 2382,2399, 2416, 2433, 2450, 2467, 2484, 2501, 2518, 2535, 2552, 2569, 2586,2603, 2620, 2637, 2654, 2671, 2688, 2705, 2722, 2739, 2773, 2790, 2824,2841, 2858, 2926, 2943, 2960, 3011, 3028, 3062, 3079, 3096, 3113, 3130,3147, 3164, 3181, 3198, 3232, 3249, 3266, and 3283; or fragmentsthereof. Such homologous sequences can comprise, for example, thesequences corresponding to the genomic sequences Alicyclobacillusacidocaldarius. In the same manner, these specific homologous sequencescan correspond to variations linked to mutations within strains ofAlicyclobacillus acidocaldarius and especially correspond totruncations, substitutions, deletions and/or additions of at least onenucleotide. As will be apparent to one of ordinary skill in the art,such homologues are easily created and identified using conventionaltechniques and publicly available computer programs such as BLAST.Accordingly, each homologue referenced above should be considered as setforth herein and fully described.

Embodiments of the invention comprise the isolated and/or purifiedpolypeptides coded for by a nucleotide sequence according to theinvention, or fragments thereof, whose sequence is represented by afragment. Amino acid sequences corresponding to the isolated and/orpurified polypeptides which can be coded for according to one of thethree possible reading frames of at least one of the sequences SEQ IDNos. 2, 19, 36, 53, 70, 87, 104, 121, 138, 155, 172, 189, 206, 223, 240,257, 274, 291, 308, 325, 359, 376, 410, 427, 444, 512, 529, 546, 597,614, 648, 665, 682, 699, 716, 733, 750, 767, 784, 818, 835, 852, 886,903, 920, 937, 954, 971, 988, 1005, 1039, 1056, 1073, 1090, 1107, 1124,1141, 1158, 1175, 1192, 1209, 1226, 1243, 1260, 1277, 1294, 1311, 1328,1345, 1362, 1379, 1396, 1413, 1430, 1447, 1464, 1481, 1498, 1515, 1532,1549, 1566, 1583, 1600, 1634, 1651, 1685, 1702, 1719, 1787, 1804, 1821,1872, 1889, 1923, 1940, 1957, 1974, 1991, 2008, 2025, 2042, 2059, 2093,2110, 2127, 2161, 2178, 2195, 2212, 2229, 2246, 2263, 2280, 2314, 2331,2348, 2365, 2382, 2399, 2416, 2433, 2450, 2467, 2484, 2501, 2518, 2535,2552, 2569, 2586, 2603, 2620, 2637, 2654, 2671, 2688, 2705, 2722, 2739,2773, 2790, 2824, 2841, 2858, 2926, 2943, 2960, 3011, 3028, 3062, 3079,3096, 3113, 3130, 3147, 3164, 3181, 3198, 3232, 3249, 3266, and 3283.

Embodiments of the invention likewise relate to the isolated and/orpurified polypeptides, characterized in that they comprise a polypeptideselected from at least one of the amino acid sequences of SEQ ID Nos. 1,18, 35, 52, 69, 86, 103, 120, 137, 154, 171, 188, 205, 222, 239, 256,273, 290, 307, 324, 358, 375, 409, 426, 443, 511, 528, 545, 596, 613,647, 664, 681, 698, 715, 732, 749, 766, 783, 817, 834, 851, 885, 902,819, 936, 953, 970, 987, 1004, 1038, 1055, 1072, 1089, 1106, 1123, 1140,1157, 1174, 1191, 1208, 1225, 1242, 1259, 1276, 1293, 1310, 1327, 1344,1361, 1378, 1395, 1412, 1429, 1446, 1463, 1480, 1497, 1514, 1531, 1548,1565, 1582, 1599, 1633, 1650, 1684, 1701, 1718, 1786, 1803, 1820, 1871,1888, 1922, 1939, 1956, 1973, 1990, 2007, 2024, 2041, 2058, 2092, 2109,2126, 2160, 2177, 2094, 2211, 2228, 2245, 2262, 2279, 2313, 2330, 2347,2364, 2381, and 23982415, 2432, 2449, 2466, 2483, 2500, 2517, 2534,2551, 2568, 2585, 2602, 2619, 2636, 2653, 2670, 2687, 2704, 2721, 2738,2772, 2789, 2823, 2840, 2857, 2925, 2942, 2959, 3010, 3027, 3061, 3078,3095, 3112, 3129, 3146, 3163, 3180, 3197, 3231, 3248, 3265, and 3282; orone of their fragments.

Among the isolated and/or purified polypeptides, according toembodiments of the invention, are the isolated and/or purifiedpolypeptides of amino acid sequence SEQ ID Nos. 8-12, 25-29, 42-46,59-63, 76-80, 93-97, 110-114, 127-131, 144-148, 161-165, 178-182,195-199, 212-216, 229-233, 246-250, 263-267, 280-284, 297-301, 314-318,331-335, 348-352, 365-369, 382,-386, 399-403, 416-420, 433-437, 450-454,467-471, 484-488, 501-505, 518-522, 535-539, 552-556, 569-573, 586-590,603-607, 620-624, 637-641, 654-658, 671-675, 688-692, 705-709, 722-726,739-743, 756-760, 773-777, 790-794, 807-811, 824-828, 841-845, 858-862,875-879, 892-896, 909-913, 926-930, 943-947, 960-964, 977-981, 994-998,1011-1015, 1028-1032, 1045-1049, 1062-1066, 1079-1083, 1096-1100,1113-1117, 1130-1134, 1147-1151, 1164-1168, 1181-1185, 1198-1202,1215-1219, 1232-1236, 1249-1253, 1266-1270, 1283-1287, 1300-1304,1317-1321, 1334-1338, 1351-1355, 1368-1372, 1385-1389, 1402-1406,1419-1423, 1436-1440, 1453-1457, 1470-1474, 1487-1491, 1504-1508,1521-1525, 1538-1542, 1555-1559, 1572-1576, 1589-1593, 1606-1610,1623-1627, 1640-1644, 1657-1661, 1674-1678, 1691-1695, 1708-1712,1725-1729, 1742-1746, 1759-1763, 1776-1780, 1793-1797, 1810-1814,1827-1831, 1844-1848, 1861-1865, 1878-1882, 1895-1899, 1912-1916,1929-1933, 1946-1950, 1963-1967, 1980-1984, 1997-2001, 2014-2018,2031-2035, 2048-2052, 2065-2069, 2082-2086, 2099-2103, 2116-2120,2133-2137, 2150-2154, 2167-2171, 2184-2188, 2201-2205, 2218-2222,2235-2239, 2252-2256, 2269-2273, 2286-2290, 2303-2307, 2320-2324,2337-2341, 2354-2358, 2371-2375, 2388-2392, 2405-2409, 2422-2426,2439-2443, 2456-2460, 2473-2477, 2490-2494, 2507-2511, 2524-2528,2541-2545, 2558-2562, 2575-2579, 2592-2596, 2609-2613, 2626-2630,2643-2647, 2660-2664, 2677-2681, 2694-2698, 2711-2715, 2728-2732,2745-2749, 2762-2766, 2779-2783, 2796-2800, 2813-2817, 2830-2834,2847-2851, 2864-2868, 2881-2885, 2898-2902, 2915-2919, 2932-2936,2949-2953, 2966-2970, 2983-2987, 3000-3004, 3017-3021, 3034-3038,3051-3055, 3068-3072, 3085-3089, 3102-3106, 3119-3123, 3136-3140,3153-3157, 3170-3174, 3187-3191, 3204-3208, 3221-3225, 3238-3242,3255-3259, 3272-3276, and 3289-3293; or fragments thereof or any otherisolated and/or purified polypeptides which have a homology of at least80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%,94%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.6%, or 99.7% identity with atleast one of the sequences of SEQ ID Nos. 1, 18, 35, 52, 69, 86, 103,120, 137, 154, 171, 188, 205, 222, 239, 256, 273, 290, 307, 324, 358,375, 409, 426, 443, 511, 528, 545, 596, 613, 647, 664, 681, 698, 715,732, 749, 766, 783, 817, 834, 851, 885, 902, 819, 936, 953, 970, 987,1004, 1038, 1055, 1072, 1089, 1106, 1123, 1140, 1157, 1174, 1191, 1208,1225, 1242, 1259, 1276, 1293, 1310, 1327, 1344, 1361, 1378, 1395, 1412,1429, 1446, 1463, 1480, 1497, 1514, 1531, 1548, 1565, 1582, 1599, 1633,1650, 1684, 1701, 1718, 1786, 1803, 1820, 1871, 1888, 1922, 1939, 1956,1973, 1990, 2007, 2024, 2041, 2058, 2092, 2109, 2126, 2160, 2177, 2094,2211, 2228, 2245, 2262, 2279, 2313, 2330, 2347, 2364, 2381, 2398, 2415,2432, 2449, 2466, 2483, 2500, 2517, 2534, 2551, 2568, 2585, 2602, 2619,2636, 2653, 2670, 2687, 2704, 2721, 2738, 2772, 2789, 2823, 2840, 2857,2925, 2942, 2959, 3010, 3027, 3061, 3078, 3095, 3112, 3129, 3146, 3163,3180, 3197, 3231, 3248, 3265, and 3282; or fragments thereof. As will beapparent to one of ordinary skill in the art, such homologues are easilycreated and identified using conventional techniques and publiclyavailable computer programs such as BLAST. Accordingly, each homologuereferenced above should be considered as set forth herein and fullydescribed.

Embodiments of the invention also relate to the polypeptides,characterized in that they comprise a polypeptide selected from: a) aspecific fragment of at least 5 amino acids of a polypeptide of an aminoacid sequence according to the invention; b) a polypeptide homologous toa polypeptide such as defined in a); c) a specific biologically activefragment of a polypeptide such as defined in a) or b); and d) apolypeptide modified by a polypeptide such as defined in a), b) or c).

In the present description, the terms polypeptide, peptide and proteinare interchangeable.

In embodiments of the invention, the isolated and/or purifiedpolypeptides according to the invention may be glycosylated, pegylated,and/or otherwise post-translationally modified. In further embodiments,glycosylation, pegylation, and/or other post-translational modificationsmay occur in vivo or in vitro and/or may be performed using chemicaltechniques. In additional embodiments, any glycosylation, pegylationand/or other post-translational modifications may be N-linked orO-linked.

In embodiments of the invention any one of the isolated and/or purifiedpolypeptides according to the invention may be enzymatically orfunctionally active at temperatures at or above about 25, 30, 35, 40,45, 50, 55, 60, 65, 70, 75, 80, 85, 90, and/or 95 degrees Celsius and/ormay be enzymatically or functionally active at a pH at, below, and/orabove 8, 7, 6, 5, 4, 3, 2, 1, and/or 0. In further embodiments of theinvention, glycosylation, pegylation, and/or other posttranslationalmodification may be required for the isolated and/or purifiedpolypeptides according to the invention to be enzymatically orfunctionally active at pH at or below 8, 7, 6, 5, 4, 3, 2, 1, and/or 0or at a temperatures at or above about 25, 30, 35, 40, 45, 50, 55, 60,65, 70, 75, 80, 85, 90, and/or 95 degrees Celsius.

Aspects of the invention relate to polypeptides that are isolated orobtained by purification from natural sources, or else obtained bygenetic recombination, or alternatively by chemical synthesis and thatthey may thus contain unnatural amino acids, as will be described below.

A “polypeptide fragment” according to the embodiments of the inventionis understood as designating a polypeptide containing at least 5consecutive amino acids, preferably 10 consecutive amino acids or 15consecutive amino acids.

In the present invention, a specific polypeptide fragment is understoodas designating the consecutive polypeptide fragment coded for by aspecific fragment nucleotide sequence according to the invention.

“Homologous polypeptide” will be understood as designating thepolypeptides having, with respect to the natural polypeptide, certainmodifications such as, in particular, a deletion, addition, orsubstitution of at least one amino acid, a truncation, a prolongation, achimeric fusion, and/or a mutation. Among the homologous polypeptides,those are preferred whose amino acid sequence has at least 80% or 90%,homology with the sequences of amino acids of polypeptides according tothe invention.

“Specific homologous polypeptide” will be understood as designating thehomologous polypeptides such as defined above and having a specificfragment of polypeptide according to the invention. In the case of asubstitution, one or more consecutive or nonconsecutive amino acids arereplaced by “equivalent” amino acids. The expression “equivalent” aminoacid is directed here at designating any amino acid capable of beingsubstituted by one of the amino acids of the base structure without,however, essentially modifying the biological activities of thecorresponding peptides and such that they will be defined by thefollowing. As will be apparent to one of ordinary skill in the art, suchsubstitutions are easily created and identified using standard molecularbiology techniques and publicly available computer programs such asBLAST. Accordingly, each substitution referenced above should beconsidered as set forth herein and fully described. Examples of suchsubstitutions in the amino acid sequences SEQ ID Nos. 1, 18, 35, 52, 69,86, 103, 120, 137, 154, 171, 188, 205, 222, 239, 256, 273, 290, 307,324, 358, 375, 409, 426, 443, 511, 528, 545, 596, 613, 647, 664, 681,698, 715, 732, 749, 766, 783, 817, 834, 851, 885, 902, 819, 936, 953,970, 987, 1004, 1038, 1055, 1072, 1089, 1106, 1123, 1140, 1157, 1174,1191, 1208, 1225, 1242, 1259, 1276, 1293, 1310, 1327, 1344, 1361, 1378,1395, 1412, 1429, 1446, 1463, 1480, 1497, 1514, 1531, 1548, 1565, 1582,1599, 1633, 1650, 1684, 1701, 1718, 1786, 1803, 1820, 1871, 1888, 1922,1939, 1956, 1973, 1990, 2007, 2024, 2041, 2058, 2092, 2109, 2126, 2160,2177, 2094, 2211, 2228, 2245, 2262, 2279, 2313, 2330, 2347, 2364, 2381,2398, 2415, 2432, 2449, 2466, 2483, 2500, 2517, 2534, 2551, 2568, 2585,2602, 2619, 2636, 2653, 2670, 2687, 2704, 2721, 2738, 2772, 2789, 2823,2840, 2857, 2925, 2942, 2959, 3010, 3027, 3061, 3078, 3095, 3112, 3129,3146, 3163, 3180, 3197, 3231, 3248, 3265, and 3282 may include thoseisolated and/or purified polypeptides of amino acid sequence SEQ ID Nos.8-12, 25-29, 42-46, 59-63, 76-80, 93-97, 110-114, 127-131, 144-148,161-165, 178-182, 195-199, 212-216, 229-233, 246-250, 263-267, 280-284,297-301, 314-318, 331-335, 348-352, 365-369, 382-386, 399-403, 416-420,433-437, 450-454, 467-471, 484-488, 501-505, 518-522, 535-539, 552-556,569-573, 586-590, 603-607, 620-624, 637-641, 654-658, 671-675, 688-692,705-709, 722-726, 739-743, 756-760, 773-777, 790-794, 807-811, 824-828,841-845, 858-862, 875-879, 892-896, 909-913, 926-930, 943-947, 960-964,977-981, 994-998, 1011-1015, 1028-1032, 1045-1049, 1062-1066, 1079-1083,1096-1100, 1113-1117, 1130-1134, 1147-1151, 1164-1168, 1181-1185,1198-1202, 1215-1219, 1232-1236, 1249-1253, 1266-1270, 1283-1287,1300-1304, 1317-1321, 1334-1338, 1351-1355, 1368-1372, 1385-1389,1402-1406, 1419-1423, 1436-1440, 1453-1457, 1470-1474, 1487-1491,1504-1508, 1521-1525, 1538-1542, 1555-1559, 1572-1576, 1589-1593,1606-1610, 1623-1627, 1640-1644, 1657-1661, 1674-1678, 1691-1695,1708-1712, 1725-1729, 1742-1746, 1759-1763, 1776-1780, 1793-1797,1810-1814, 1827-1831, 1844-1848, 1861-1865, 1878-1882, 1895-1899,1912-1916, 1929-1933, 1946-1950, 1963-1967, 1980-1984, 1997-2001,2014-2018, 2031-2035, 2048-2052, 2065-2069, 2082-2086, 2099-2103,2116-2120, 2133-2137, 2150-2154, 2167-2171, 2184-2188, 2201-2205,2218-2222, 2235-2239, 2252-2256, 2269-2273, 2286-2290, 2303-2307,2320-2324, 2337-2341, 2354-2358, 2371-2375, 2388-2392, 2405-2409,2422-2426, 2439-2443, 2456-2460, 2473-2477, 2490-2494, 2507-2511,2524-2528, 2541-2545, 2558-2562, 2575-2579, 2592-2596, 2609-2613,2626-2630, 2643-2647, 2660-2664, 2677-2681, 2694-2698, 2711-2715,2728-2732, 2745-2749, 2762-2766, 2779-2783, 2796-2800, 2813-2817,2830-2834, 2847-2851, 2864-2868, 2881-2885, 2898-2902, 2915-2919,2932-2936, 2949-2953, 2966-2970, 2983-2987, 3000-3004, 3017-3021,3034-3038, 3051-3055, 3068-3072, 3085-3089, 3102-3106, 3119-3123,3136-3140, 3153-3157, 3170-3174, 3187-3191, 3204-3208, 3221-3225,3238-3242, 3255-3259, 3272-3276, and 3289-3293. These equivalent aminoacids may be determined either by depending on their structural homologywith the amino acids which they substitute, or on results of comparativetests of biological activity between the different polypeptides, whichare capable of being carried out.

By way of nonlimiting example, the possibilities of substitutionscapable of being carried out without resulting in an extensivemodification of the biological activity of the corresponding modifiedpolypeptides will be mentioned, the replacement, for example, of leucineby valine or isoleucine, of aspartic acid by glutamic acid, of glutamineby asparagine, of arginine by lysine etc., the reverse substitutionsnaturally being envisageable under the same conditions.

In a further embodiment, substitutions are limited to substitutions inamino acids not conserved among other proteins which have similaridentified enzymatic activity. For example, one of ordinary skill in theart may align proteins of the same function in similar organisms anddetermine which amino acids are generally conserved among proteins ofthat function. One example of a program that may be used to generatesuch alignments is worldwideweb.charite.de/bioinf/strap/ in conjunctionwith the databases provided by the NCBI. Examples of such polypeptidesmay include, but are not limited to, those found in amino acid sequenceSEQ ID Nos. 8-12, 25-29, 42-46, 59-63, 76-80, 93-97, 110-114, 127-131,144-148, 161-165, 178-182, 195-199, 212-216, 229-233, 246-250, 263-267,280-284, 297-301, 314-318, 331-335, 348-352, 365-369, 382-386, 399-403,416-420, 433-437, 450-454, 467-471, 484-488, 501-505, 518-522, 535-539,552-556, 569-573, 586-590, 603-607, 620-624, 637-641, 654-658, 671-675,688-692, 705-709, 722-726, 739-743, 756-760, 773-777, 790-794, 807-811,824-828, 841-845, 858-862, 875-879, 892-896, 909-913, 926-930, 943-947,960-964, 977-981, 994-998, 1011-1015, 1028-1032, 1045-1049, 1062-1066,1079-1083, 1096-1100, 1113-1117, 1130-1134, 1147-1151, 1164-1168,1181-1185, 1198-1202, 1215-1219, 1232-1236, 1249-1253, 1266-1270,1283-1287, 1300-1304, 1317-1321, 1334-1338, 1351-1355, 1368-1372,1385-1389, 1402-1406, 1419-1423, 1436-1440, 1453-1457, 1470-1474,1487-1491, 1504-1508, 1521-1525, 1538-1542, 1555-1559, 1572-1576,1589-1593, 1606-1610, 1623-1627, 1640-1644, 1657-1661, 1674-1678,1691-1695, 1708-1712, 1725-1729, 1742-1746, 1759-1763, 1776-1780,1793-1797, 1810-1814, 1827-1831, 1844-1848, 1861-1865, 1878-1882,1895-1899, 1912-1916, 1929-1933, 1946-1950, 1963-1967, 1980-1984,1997-2001, 2014-2018, 2031-2035, 2048-2052, 2065-2069, 2082-2086,2099-2103, 2116-2120, 2133-2137, 2150-2154, 2167-2171, 2184-2188,2201-2205, 2218-2222, 2235-2239, 2252-2256, 2269-2273, 2286-2290,2303-2307, 2320-2324, 2337-2341, 2354-2358, 2371-2375, 2388-2392,2405-2409, 2422-2426, 2439-2443, 2456-2460, 2473-2477, 2490-2494,2507-2511, 2524-2528, 2541-2545, 2558-2562, 2575-2579, 2592-2596,2609-2613, 2626-2630, 2643-2647, 2660-2664, 2677-2681, 2694-2698,2711-2715, 2728-2732, 2745-2749, 2762-2766, 2779-2783, 2796-2800,2813-2817, 2830-2834, 2847-2851, 2864-2868, 2881-2885, 2898-2902,2915-2919, 2932-2936, 2949-2953, 2966-2970, 2983-2987, 3000-3004,3017-3021, 3034-3038, 3051-3055, 3068-3072, 3085-3089, 3102-3106,3119-3123, 3136-3140, 3153-3157, 3170-3174, 3187-3191, 3204-3208,3221-3225, 3238-3242, 3255-3259, 3272-3276, and 3289-3293.

Thus, according to one embodiment of the invention, substitutions ormutation may be made at positions that are generally conserved amongproteins of that function. In a further embodiment, nucleic acidsequences may be mutated or substituted such that the amino acid theycode for is unchanged (degenerate substitutions and/mutations) and/ormutated or substituted such that any resulting amino acid substitutionsor mutation are made at positions that are generally conserved amongproteins of that function. Examples of such nucleic acid sequences mayinclude, but are not limited to, those found in are the nucleotidesequences of SEQ ID Nos. 13-17, 30-34, 47-51, 64-68, 81-85, 98-102,115-119, 132-136, 149-153, 166-170, 183-187, 200-204, 217-221, 234-238,251-255, 268-272, 285-289, 302-306, 319-323, 336-340, 353-357, 370-374,387-391, 404-408, 421-425, 438-442, 455-459, 472-476, 489-493, 506-510,523-527, 540-544, 557-561, 574-578, 591-595, 608-612, 625-629, 642-646,659-663, 676-680, 693-697, 710-714, 727-731, 744-748, 761-765, 778-782,795-799, 812-816, 829-833, 846-850, 863-867, 880-884, 897-901, 914-918,931-935, 948-952, 965-969, 982-986, 999-1003, 1016-1020, 1033-1037,1050-1054, 1067-1071, 1084-1088, 1101-1105, 1118-1122, 1135-1139,1152-1156, 1169-1173, 1186-1190, 1203-1207, 1220-1224, 1237-1241,1254-1258, 1271-1275, 1288-1292, 1305-1309, 1322-1326, 1339-1343,1356-1360, 1373-1377, 1390-1394, 1407-1411, 1424-1428, 1441-1445,1458-1462, 1475-1479, 1492-1496, 1509-1513, 1526-1530, 1543-1547,1560-1564, 1577-1581, 1594-1598, 1611-1615, 1628-1632, 1645-1649,1662-1666, 1679-1683, 1696-1700, 1713-1717, 1730-1734, 1747-1751,1764-1768, 1781-1785, 1798-1802, 1815-1819, 1832-1836, 1849-1853,1866-1870, 1883-1887, 1900-1904, 1917-1921, 1934-1938, 1951-1955,1968-1972, 1985-1989, 2002-2006, 2019-2023, 2036-2040, 2053-2057,2070-2074, 2087-2091, 2104-2108, 2121-2125, 2138-2142, 2155-2159,2172-2176, 2189-2193, 2206-2210, 2223-2227, 2240-2244, 2257-2261,2274-2278, 2291-2295, 2308-2312, 2325-2329, 2342-2346, 2359-2363,2376-2380, 2393-2397, 2410-2414, 2427-2431, 2444-2448, 2461-2465,2478-2482, 2495-2499, 2512-2516, 2529-2533, 2546-2550, 2563-2567,2580-2584, 2597-2601, 2614-2618, 2631-2635, 2648-2652, 2665-2669,2682-2686, 2699-2703, 2716-2720, 2733-2737, 2750-2754, 2767-2771,2784-2788, 2801-2805, 2818-2822, 2835-2839, 2852-2856, 2869-2873,2886-2890, 2903-2907, 2920-2924, 2937-2941, 2954-2958, 2971-2975,2988-2992, 3005-3009, 3022-3026, 3039-3043, 3056-3060, 3073-3077,3090-3094, 3107-3111, 3124-3128, 3141-3145, 3158-3162, 3175-3179,3192-3196, 3209-3213, 3226-3230, 3243-3247, 3260-3264, 3277-3281, and3294-3298; or fragments thereof.

The specific homologous polypeptides likewise correspond to polypeptidescoded for by the specific homologous nucleotide sequences such asdefined above and thus comprise in the present definition thepolypeptides which are mutated or correspond to variants which can existin Alicyclobacillus acidocaldarius, and which especially correspond totruncations, substitutions, deletions, and/or additions of at least oneamino acid residue.

“Specific biologically active fragment of a polypeptide” according to anembodiment of the invention will be understood in particular asdesignating a specific polypeptide fragment, such as defined above,having at least one of the characteristics of polypeptides according tothe invention. In certain embodiments the peptide is capable of behavingas at least one of the types of proteins outlined in Table 1.

The polypeptide fragments according to embodiments of the invention cancorrespond to isolated or purified fragments naturally present inAlicyclobacillus acidocaldarius or correspond to fragments which can beobtained by cleavage of said polypeptide by a proteolytic enzyme, suchas trypsin or chymotrypsin or collagenase, or by a chemical reagent,such as cyanogen bromide (CNBr). Such polypeptide fragments can likewisejust as easily be prepared by chemical synthesis, from hosts transformedby an expression vector according to the invention containing a nucleicacid allowing the expression of said fragments, placed under the controlof appropriate regulation and/or expression elements.

“Modified polypeptide” of a polypeptide according to an embodiment ofthe invention is understood as designating a polypeptide obtained bygenetic recombination or by chemical synthesis as will be describedbelow, having at least one modification with respect to the normalsequence. These modifications may or may not be able to bear on aminoacids at the origin of specificity, and/or of activity, or at the originof the structural conformation, localization, and of the capacity ofmembrane insertion of the polypeptide according to the invention. Itwill thus be possible to create polypeptides of equivalent, increased,or decreased activity, and of equivalent, narrower, or widerspecificity. Among the modified polypeptides, it is necessary to mentionthe polypeptides in which up to 5 or more amino acids can be modified,truncated at the N- or C-terminal end, or even deleted or added.

The methods allowing said modulations on eukaryotic or prokaryotic cellsto be demonstrated are well known to the person of ordinary skill in theart. It is likewise well understood that it will be possible to use thenucleotide sequences coding for said modified polypeptides for saidmodulations, for example through vectors according to the invention anddescribed below.

The preceding modified polypeptides can be obtained by usingcombinatorial chemistry, in which it is possible to systematically varyparts of the polypeptide before testing them on models, cell cultures ormicroorganisms for example, to select the compounds which are mostactive or have the properties sought.

Chemical synthesis likewise has the advantage of being able to usenonnatural amino acids, or nonpeptide bonds.

Thus, in order to improve the duration of life of the polypeptidesaccording to the invention, it may be of interest to use nonnaturalamino acids, for example in D form, or else amino acid analogs,especially sulfur-containing forms, for example.

Finally, it will be possible to integrate the structure of thepolypeptides according to the invention, its specific or modifiedhomologous forms, into chemical structures of polypeptide type orothers. Thus, it may be of interest to provide at the N- and C-terminalends molecules not recognized by proteases.

The nucleotide sequences coding for a polypeptide according to theinvention are likewise part of the invention.

The invention likewise relates to nucleotide sequences utilizable as aprimer or probe, characterized in that said sequences are selected fromthe nucleotide sequences according to the invention.

It is well understood that the present invention, in variousembodiments, likewise relates to specific polypeptides ofAlicyclobacillus acidocaldarius, coded for by nucleotide sequences,capable of being obtained by purification from natural polypeptides, bygenetic recombination or by chemical synthesis by procedures well knownto the person skilled in the art and such as described in particularbelow. In the same manner, the labeled or unlabeled mono- or polyclonalantibodies directed against said specific polypeptides coded for by saidnucleotide sequences are also encompassed by the invention.

Embodiments of the invention additionally relate to the use of anucleotide sequence according to the invention as a primer or probe forthe detection and/or the amplification of nucleic acid sequences.

The nucleotide sequences according to embodiments of the invention canthus be used to amplify nucleotide sequences, especially by the PCRtechnique (polymerase chain reaction) (Erlich, 1989; Innis et al., 1990;Rolfs et al., 1991; and White et al., 1997).

These oligodeoxyribonucleotide or oligoribonucleotide primersadvantageously have a length of at least 8 nucleotides, preferably of atleast 12 nucleotides, and even more preferentially at least 20nucleotides.

Other amplification techniques of the target nucleic acid can beadvantageously employed as alternatives to PCR.

The nucleotide sequences of the invention, in particular the primersaccording to the invention, can likewise be employed in other proceduresof amplification of a target nucleic acid, such as: the TAS technique(Transcription-based Amplification System), described by Kwoh et al. in1989; the 3SR technique (Self-Sustained Sequence Replication), describedby Guatelli et al. in 1990; the NASBA technique (Nucleic Acid SequenceBased Amplification), described by Kievitis et al. in 1991; the SDAtechnique (Strand Displacement Amplification) (Walker et al., 1992); theTMA technique (Transcription Mediated Amplification).

The polynucleotides of the invention can also be employed in techniquesof amplification or of modification of the nucleic acid serving as aprobe, such as: the LCR technique (Ligase Chain Reaction), described byLandegren et al. in 1988 and improved by Barany et al. in 1991, whichemploys a thermostable ligase; the RCR technique (Repair ChainReaction), described by Segev in 1992; the CPR technique (Cycling ProbeReaction), described by Duck et al. in 1990; the amplification techniquewith Q-beta replicase, described by Miele et al. in 1983 and especiallyimproved by Chu et al. in 1986, Lizardi et al. in 1988, then by Burg etal. as well as by Stone et al. in 1996.

In the case where the target polynucleotide to be detected is possiblyan RNA, for example an mRNA, it will be possible to use, prior to theemployment of an amplification reaction with the aid of at least oneprimer according to the invention or to the employment of a detectionprocedure with the aid of at least one probe of the invention, an enzymeof reverse transcriptase type in order to obtain a cDNA from the RNAcontained in the biological sample. The cDNA obtained will thus serve asa target for the primer(s) or the probe(s) employed in the amplificationor detection procedure according to the invention.

The detection probe will be chosen in such a manner that it hybridizeswith the target sequence or the amplicon generated from the targetsequence. By way of sequence, such a probe will advantageously have asequence of at least 12 nucleotides, in particular of at least 20nucleotides, and preferably of at least 100 nucleotides.

Embodiments of the invention also comprise the nucleotide sequencesutilizable as a probe or primer according to the invention,characterized in that they are labeled with a radioactive compound orwith a nonradioactive compound.

The unlabeled nucleotide sequences can be used directly as probes orprimers, although the sequences are generally labeled with a radioactiveisotope (³²P, ³⁵S, ³H, ¹²⁵I) or with a nonradioactive molecule (biotin,acetylaminofluorene, digoxigenin, 5-bromodeoxyuridine, fluorescein) toobtain probes which are utilizable for numerous applications.

Examples of nonradioactive labeling of nucleotide sequences aredescribed, for example, in French Patent No. 78.10975 or by Urdea et al.or by Sanchez-Pescador et al. in 1988.

In the latter case, it will also be possible to use one of the labelingmethods described in patents FR-2 422 956 and FR-2 518 755.

The hybridization technique can be carried out in various manners(Matthews et al., 1988). The most general method consists inimmobilizing the nucleic acid extract of cells on a support (such asnitrocellulose, nylon, polystyrene) and in incubating, underwell-defined conditions, the immobilized target nucleic acid with theprobe. After hybridization, the excess of probe is eliminated and thehybrid molecules formed are detected by the appropriate method(measurement of the radioactivity, of the fluorescence or of theenzymatic activity linked to the probe).

The invention, in various embodiments, likewise comprises the nucleotidesequences according to the invention, characterized in that they areimmobilized on a support, covalently or noncovalently.

According to another advantageous mode of employing nucleotide sequencesaccording to the invention, the latter can be used immobilized on asupport and can thus serve to capture, by specific hybridization, thetarget nucleic acid obtained from the biological sample to be tested. Ifnecessary, the solid support is separated from the sample and thehybridization complex formed between said capture probe and the targetnucleic acid is then detected with the aid of a second probe, aso-called detection probe, labeled with an easily detectable element.

Another aspect of the present invention is a vector for the cloningand/or expression of a sequence, characterized in that it contains anucleotide sequence according to the invention.

The vectors according to the invention, characterized in that theycontain the elements allowing the integration, expression and/or thesecretion of said nucleotide sequences in a determined host cell, arelikewise part of the invention.

The vector may then contain a promoter, signals of initiation andtermination of translation, as well as appropriate regions of regulationof transcription. It may be able to be maintained stably in the hostcell and can optionally have particular signals specifying the secretionof the translated protein. These different elements may be chosen as afunction of the host cell used. To this end, the nucleotide sequencesaccording to the invention may be inserted into autonomous replicationvectors within the chosen host, or integrated vectors of the chosenhost.

Such vectors will be prepared according to the methods currently used bythe person skilled in the art, and it will be possible to introduce theresulting vectors into an appropriate host by standard methods, such as,for example, lipofection, electroporation, conjugation, and thermalshock.

The vectors according to the invention are, for example, vectors ofplasmid or viral origin. One example of a vector for the expression ofpolypeptides of the invention is baculovirus.

These vectors are useful for transforming host cells in order to cloneor to express the nucleotide sequences of the invention.

The invention likewise comprises the host cells transformed by a vectoraccording to the invention.

These cells can be obtained by the introduction into host cells of anucleotide sequence inserted into a vector such as defined above, thenthe culturing of said cells under conditions allowing the replicationand/or expression of the transfected nucleotide sequence.

The host cell can be selected from prokaryotic or eukaryotic systems,such as, for example, bacterial cells (Olins and Lee, 1993), butlikewise yeast cells (Buckholz, 1993), as well as plants cells, such asArabidopsis sp., and animal cells, in particular the cultures ofmammalian cells (Edwards and Aruffo, 1993), for example, Chinese hamsterovary (CHO) cells, but likewise the cells of insects in which it ispossible to use procedures employing baculoviruses, for example sf9insect cells (Luckow, 1993).

Embodiments of the invention likewise relate to organisms comprising oneof such transformed cells according to the invention.

The obtainment of transgenic organisms according to the inventionexpressing one or more of the coding sequences of Alicyclobacillusacidocaldarius or part of the coding seqeunces may be carried out in,for example, rats, mice, or rabbits according to methods well known tothe person skilled in the art, such as by viral or nonviraltransfections. It will be possible to obtain the transgenic organismsexpressing one or more of such coding sequences by transfection ofmultiple copies of such coding sequences under the control of a strongpromoter of ubiquitous nature, or selective for one type of tissue. Itwill likewise be possible to obtain the transgenic organisms byhomologous recombination in embryonic cell strains, transfer of thesecell strains to embryos, selection of the affected chimeras at the levelof the reproductive lines, and growth of said chimeras.

The transformed cells as well as the transgenic organisms according tothe invention are utilizable in procedures for preparation ofrecombinant polypeptides.

It is today possible to produce recombinant polypeptides in relativelylarge quantity by genetic engineering using the cells transformed byexpression vectors according to the invention or using transgenicorganisms according to the invention.

The procedures for preparation of a polypeptide of the invention inrecombinant form, characterized in that they employ a vector and/or acell transformed by a vector according to the invention and/or atransgenic organism comprising one of said transformed cells accordingto the invention are themselves comprised in the present invention.

As used herein, “transformation” and “transformed” relate to theintroduction of nucleic acids into a cell, whether prokaryotic oreukaryotic. Further, “transformation” and “transformed,” as used herein,need not relate to growth control or growth deregulation.

Among the procedures for preparation of a polypeptide of the inventionin recombinant form, the preparation procedures employing a vector,and/or a cell transformed by the vector and/or a transgenic organismcomprising one of the transformed cells, containing a nucleotidesequence according to the invention coding for a polypeptide ofAlicyclobacillus acidocaldarius.

A variant according to the invention may consist of producing arecombinant polypeptide fused to a “carrier” protein (chimeric protein).The advantage of this system is that it may allow stabilization ofand/or a decrease in the proteolysis of the recombinant product, anincrease in the solubility in the course of renaturation in vitro and/ora simplification of the purification when the fusion partner has anaffinity for a specific ligand.

More particularly, the invention relates to a procedure for preparationof a polypeptide of the invention comprising the following steps: a)culture of transformed cells under conditions allowing the expression ofa recombinant polypeptide of nucleotide sequence according to theinvention; b) if need be, recovery of the recombinant polypeptide.

When the procedure for preparation of a polypeptide of the inventionemploys a transgenic organism according to the invention, therecombinant polypeptide is then extracted from said organism.

The invention also relates to a polypeptide which is capable of beingobtained by a procedure of the invention such as described previously.

The invention also comprises a procedure for preparation of a syntheticpolypeptide, characterized in that it uses a sequence of amino acids ofpolypeptides according to the invention.

The invention likewise relates to a synthetic polypeptide obtained by aprocedure according to the invention.

The polypeptides according to the invention can likewise be prepared bytechniques which are conventional in the field of the synthesis ofpeptides. This synthesis can be carried out in homogeneous solution orin solid phase.

For example, recourse can be made to the technique of synthesis inhomogeneous solution described by Houben-Weyl in 1974.

This method of synthesis consists in successively condensing, two bytwo, the successive amino acids in the order required, or in condensingamino acids and fragments formed previously and already containingseveral amino acids in the appropriate order, or alternatively severalfragments previously prepared in this way, it being understood that itwill be necessary to protect beforehand all the reactive functionscarried by these amino acids or fragments, with the exception of aminefunctions of one and carboxyls of the other or vice-versa, which mustnormally be involved in the formation of peptide bonds, especially afteractivation of the carboxyl function, according to the methods well knownin the synthesis of peptides.

Recourse may also be made to the technique described by Merrifield.

To make a peptide chain according to the Merrifield procedure, recourseis made to a very porous polymeric resin, on which is immobilized thefirst C-terminal amino acid of the chain. This amino acid is immobilizedon a resin through its carboxyl group and its amine function isprotected. The amino acids which are going to form the peptide chain arethus immobilized, one after the other, on the amino group, which isdeprotected beforehand each time, of the portion of the peptide chainalready formed, and which is attached to the resin. When the whole ofthe desired peptide chain has been formed, the protective groups of thedifferent amino acids forming the peptide chain are eliminated and thepeptide is detached from the resin with the aid of an acid.

The invention additionally relates to hybrid polypeptides having atleast one polypeptide according to the invention, and a sequence of apolypeptide capable of inducing an immune response in man or animals.

Advantageously, the antigenic determinant is such that it is capable ofinducing a humoral and/or cellular response.

It will be possible for such a determinant to comprise a polypeptideaccording to the invention in glycosylated, pegylated, and/or otherwisepost-translationally modified form used with a view to obtainingimmunogenic compositions capable of inducing the synthesis of antibodiesdirected against multiple epitopes.

These hybrid molecules can be formed, in part, of a polypeptide carriermolecule or of fragments thereof according to the invention, associatedwith a possibly immunogenic part, in particular an epitope of thediphtheria toxin, the tetanus toxin, a surface antigen of the hepatitisB virus (patent FR 79 21811), the VP1 antigen of the poliomyelitis virusor any other viral or bacterial toxin or antigen.

The procedures for synthesis of hybrid molecules encompass the methodsused in genetic engineering for constructing hybrid nucleotide sequencescoding for the polypeptide sequences sought. It will be possible, forexample, to refer advantageously to the technique for obtainment ofgenes coding for fusion proteins described by Minton in 1984.

The hybrid nucleotide sequences coding for a hybrid polypeptide as wellas the hybrid polypeptides according to the invention characterized inthat they are recombinant polypeptides obtained by the expression of thehybrid nucleotide sequences are likewise part of the invention.

The invention likewise comprises the vectors characterized in that theycontain one of the hybrid nucleotide sequences. The host cellstransformed by the vectors, the transgenic organisms comprising one ofsaid transformed cells as well as the procedures for preparation ofrecombinant polypeptides using the vectors, the transformed cells and/orsaid transgenic organisms are, of course, likewise part of theinvention.

The polypeptides according to the invention, the antibodies according tothe invention described below and the nucleotide sequences according tothe invention can advantageously be employed in procedures for thedetection and/or identification of Alicyclobacillus acidocaldarius, in asample capable of containing them. These procedures, according to thespecificity of the polypeptides, the antibodies and the nucleotidesequences according to the invention which will be used, will inparticular be able to detect and/or to identify Alicyclobacillusacidocaldarius.

The polypeptides according to the invention can advantageously beemployed in a procedure for the detection and/or the identification ofAlicyclobacillus acidocaldarius in a sample capable of containing them,characterized in that it comprises the following steps: a) contacting ofthis sample with a polypeptide or one of its fragments according to theinvention (under conditions allowing an immunological reaction betweensaid polypeptide and the antibodies possibly present in the biologicalsample); b) demonstration of the antigen-antibody complexes possiblyformed.

Any conventional procedure can be employed for carrying out such adetection of the antigen-antibody complexes possibly formed.

By way of nonlimiting example, one method brings into playimmunoenzymatic processes according to the ELISA technique, byimmunofluorescence, or radioimmunological processes (RIA) or theirequivalent.

Thus, the invention likewise relates to the polypeptides according tothe invention, labeled with the aid of an adequate label, such as, ofthe enzymatic, fluorescent or radioactive type.

Such methods comprise, for example, the following acts: deposition ofdetermined quantities of a polypeptide composition according to theinvention in the wells of a microtiter plate, introduction into thewells of increasing dilutions of serum, or of a biological sample otherthan that defined previously, having to be analyzed, incubation of themicrotiter plate, introduction into the wells of the microtiter plate oflabeled antibodies directed against pig immunoglobulins, the labeling ofthese antibodies having been carried out with the aid of an enzymeselected from those which are capable of hydrolyzing a substrate bymodifying the absorption of the radiation of the latter, at least at adetermined wavelength, for example at 550 nm, detection, by comparisonwith a control test, of the quantity of hydrolyzed substrate.

The polypeptides according to the invention allow monoclonal orpolyclonal antibodies to be prepared which are characterized in thatthey specifically recognize the polypeptides according to the invention.It will advantageously be possible to prepare the monoclonal antibodiesfrom hybridomas according to the technique described by Kohler andMilstein in 1975. It will be possible to prepare the polyclonalantibodies, for example, by immunization of an animal, in particular amouse, with a polypeptide or a DNA, according to the invention,associated with an adjuvant of the immune response, and thenpurification of the specific antibodies contained in the serum of theimmunized animals on an affinity column on which the polypeptide whichhas served as an antigen has previously been immobilized. The polyclonalantibodies according to the invention can also be prepared bypurification, on an affinity column on which a polypeptide according tothe invention has previously been immobilized, of the antibodiescontained in the serum of an animal immunologically challenged byAlicyclobacillus acidocaldarius, or a polypeptide or fragment accordingto the invention.

The invention likewise relates to mono- or polyclonal antibodies ortheir fragments, or chimeric antibodies, characterized in that they arecapable of specifically recognizing a polypeptide according to theinvention.

It will likewise be possible for the antibodies of the invention to belabeled in the same manner as described previously for the nucleicprobes of the invention, such as a labeling of enzymatic, fluorescent orradioactive type.

The invention is additionally directed at a procedure for the detectionand/or identification of Alicyclobacillus acidocaldarius in a sample,characterized in that it comprises the following steps: a) contacting ofthe sample with a mono- or polyclonal antibody according to theinvention (under conditions allowing an immunological reaction betweensaid antibodies and the polypeptides of Alicyclobacillus acidocaldariuspossibly present in the biological sample); b) demonstration of theantigen-antibody complex possibly formed.

The present invention likewise relates to a procedure for the detectionand/or the identification of Alicyclobacillus acidocaldarius in asample, characterized in that it employs a nucleotide sequence accordingto the invention.

More particularly, the invention relates to a procedure for thedetection and/or the identification of Alicyclobacillus acidocaldariusin a sample, characterized in that it contains the following steps: a)if need be, isolation of the DNA from the sample to be analyzed; b)specific amplification of the DNA of the sample with the aid of at leastone primer, or a pair of primers, according to the invention; c)demonstration of the amplification products.

These can be detected, for example, by the technique of molecularhybridization utilizing a nucleic probe according to the invention. Thisprobe will advantageously be labeled with a nonradioactive (cold probe)or radioactive isotope.

For the purposes of the present invention, “DNA of the biologicalsample” or “DNA contained in the biological sample” will be understoodas meaning either the DNA present in the biological sample considered,or possibly the cDNA obtained after the action of an enzyme of reversetranscriptase type on the RNA present in said biological sample.

A further embodiment of the invention comprises a method, characterizedin that it comprises the following acts: a) contacting of a nucleotideprobe according to the invention with a biological sample, the DNAcontained in the biological sample having, if need be, previously beenmade accessible to hybridization under conditions allowing thehybridization of the probe with the DNA of the sample; b) demonstrationof the hybrid formed between the nucleotide probe and the DNA of thebiological sample.

The present invention also relates to a procedure according to theinvention, characterized in that it comprises the following acts: a)contacting of a nucleotide probe immobilized on a support according tothe invention with a biological sample, the DNA of the sample having, ifneed be, previously been made accessible to hybridization, underconditions allowing the hybridization of the probe with the DNA of thesample; b) contacting of the hybrid formed between the nucleotide probeimmobilized on a support and the DNA contained in the biological sample,if need be after elimination of the DNA of the biological sample whichhas not hybridized with the probe, with a nucleotide probe labeledaccording to the invention; c) demonstration of the novel hybrid formedin act b).

According to an advantageous embodiment of the procedure for detectionand/or identification defined previously, this is characterized in that,prior to act a), the DNA of the biological sample is first amplifiedwith the aid of at least one primer according to the invention.Embodiments of methods include methods of altering recombination insideor outside of a cell, the methods comprising: providing a recombinant,purified, and/or isolated nucleotide sequence comprising a nucleotidesequence selected from the group consisting of a nucleotide sequenceshaving at least 90% sequence identity to at least one of the sequencesof SEQ ID Nos. 2, 19, 36, 53, 70, 87, 104, 121, 138, 155, 172, 189, 206,223, 240, 257, 274, 291, 308, 325, 359, 376, 410, 427, 444, 512, 529,546, 597, 614, 648, 665, 682, 699, 716, 733, 750, 767, 784, 818, 835,852, 886, 903, 920, 937, 954, 971, 988, 1005, 1039, 1056, 1073, 1090,1107, 1124, 1141, 1158, 1175, 1192, 1209, 1226, 1243, 1260, 1277, 1294,1311, 1328, 1345, 1362, 1379, 1396, 1413, 1430, 1447, 1464, 1481, 1498,1515, 1532, 1549, 1566, 1583, 1600, 1634, 1651, 1685, 1702, 1719, 1787,1804, 1821, 1872, 1889, 1923, 1940, 1957, 1974, 1991, 2008, 2025, 2042,2059, 2093, 2110, 2127, 2161, 2178, 2195, 2212, 2229, 2246, 2263, 2280,2314, 2331, 2348, 2365, 2382, 2399, 2416, 2433, 2450, 2467, 2484, 2501,2518, 2535, 2552, 2569, 2586, 2603, 2620, 2637, 2654, 2671, 2688, 2705,2722, 2739, 2773, 2790, 2824, 2841, 2858, 2926, 2943, 2960, 3011, 3028,3062, 3079, 3096, 3113, 3130, 3147, 3164, 3181, 3198, 3232, 3249, 3266,and 3283; and/or a recombinant, purified, and/or isolated polypeptideselected from the group consisting of a polypeptide having at least 90%sequence identity to at least one of the sequences of SEQ ID Nos. 1, 18,35, 52, 69, 86, 103, 120, 137, 154, 171, 188, 205, 222, 239, 256, 273,290, 307, 324, 358, 375, 409, 426, 443, 511, 528, 545, 596, 613, 647,664, 681, 698, 715, 732, 749, 766, 783, 817, 834, 851, 885, 902, 819,936, 953, 970, 987, 1004, 1038, 1055, 1072, 1089, 1106, 1123, 1140,1157, 1174, 1191, 1208, 1225, 1242, 1259, 1276, 1293, 1310, 1327, 1344,1361, 1378, 1395, 1412, 1429, 1446, 1463, 1480, 1497, 1514, 1531, 1548,1565, 1582, 1599, 1633, 1650, 1684, 1701, 1718, 1786, 1803, 1820, 1871,1888, 1922, 1939, 1956, 1973, 1990, 2007, 2024, 2041, 2058, 2092, 2109,2126, 2160, 2177, 2094, 2211, 2228, 2245, 2262, 2279, 2313, 2330, 2347,2364, 2381, 2398, 2415, 2432, 2449, 2466, 2483, 2500, 2517, 2534, 2551,2568, 2585, 2602, 2619, 2636, 2653, 2670, 2687, 2704, 2721, 2738, 2772,2789, 2823, 2840, 2857, 2925, 2942, 2959, 3010, 3027, 3061, 3078, 3095,3112, 3129, 3146, 3163, 3180, 3197, 3231, 3248, 3265, and 3282 to anucleotide sequence with which recombination event is desired.

Further embodiments of methods include placing a cell producing orencoding a recombinant, purified, and/or isolated nucleotide sequencecomprising a nucleotide sequence selected from the group consisting of anucleotide sequences having at least 90% sequence identity to at leastone of the sequences of SEQ ID Nos. 2, 19, 36, 53, 70, 87, 104, 121,138, 155, 172, 189, 206, 223, 240, 257, 274, 291, 308, 325, 359, 376,410, 427, 444, 512, 529, 546, 597, 614, 648, 665, 682, 699, 716, 733,750, 767, 784, 818, 835, 852, 886, 903, 920, 937, 954, 971, 988, 1005,1039, 1056, 1073, 1090, 1107, 1124, 1141, 1158, 1175, 1192, 1209, 1226,1243, 1260, 1277, 1294, 1311, 1328, 1345, 1362, 1379, 1396, 1413, 1430,1447, 1464, 1481, 1498, 1515, 1532, 1549, 1566, 1583, 1600, 1634, 1651,1685, 1702, 1719, 1787, 1804, 1821, 1872, 1889, 1923, 1940, 1957, 1974,1991, 2008, 2025, 2042, 2059, 2093, 2110, 2127, 2161, 2178, 2195, 2212,2229, 2246, 2263, 2280, 2314, 2331, 2348, 2365, 2382, 2399, 2416, 2433,2450, 2467, 2484, 2501, 2518, 2535, 2552, 2569, 2586, 2603, 2620, 2637,2654, 2671, 2688, 2705, 2722, 2739, 2773, 2790, 2824, 2841, 2858, 2926,2943, 2960, 3011, 3028, 3062, 3079, 3096, 3113, 3130, 3147, 3164, 3181,3198, 3232, 3249, 3266, and 3283; and/or a recombinant, purified, and/orisolated polypeptide selected from the group consisting of a polypeptidehaving at least 90% sequence identity to at least one of the sequencesof SEQ ID Nos. 1, 18, 35, 52, 69, 86, 103, 120, 137, 154, 171, 188, 205,222, 239, 256, 273, 290, 307, 324, 358, 375, 409, 426, 443, 511, 528,545, 596, 613, 647, 664, 681, 698, 715, 732, 749, 766, 783, 817, 834,851, 885, 902, 819, 936, 953, 970, 987, 1004, 1038, 1055, 1072, 1089,1106, 1123, 1140, 1157, 1174, 1191, 1208, 1225, 1242, 1259, 1276, 1293,1310, 1327, 1344, 1361, 1378, 1395, 1412, 1429, 1446, 1463, 1480, 1497,1514, 1531, 1548, 1565, 1582, 1599, 1633, 1650, 1684, 1701, 1718, 1786,1803, 1820, 1871, 1888, 1922, 1939, 1956, 1973, 1990, 2007, 2024, 2041,2058, 2092, 2109, 2126, 2160, 2177, 2094, 2211, 2228, 2245, 2262, 2279,2313, 2330, 2347, 2364, 2381, 2398, 2415, 2432, 2449, 2466, 2483, 2500,2517, 2534, 2551, 2568, 2585, 2602, 2619, 2636, 2653, 2670, 2687, 2704,2721, 2738, 2772, 2789, 2823, 2840, 2857, 2925, 2942, 2959, 3010, 3027,3061, 3078, 3095, 3112, 3129, 3146, 3163, 3180, 3197, 3231, 3248, 3265,and 3282 in a environment comprising temperatures at or above about 25,30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, and/or 95 degreesCelsius and/or a pH at, below, and/or above 8, 7, 6, 5, 4, 3, 2, 1,and/or 0

The present invention provides cells that have been geneticallymanipulated to have an altered capacity to produce expressed proteins.In particular, the present invention relates to Gram-positivemicroorganisms, such as Bacillus species having enhanced expression of aprotein of interest, wherein one or more chromosomal genes have beeninactivated, and/or wherein one or more chromosomal genes have beendeleted from the Bacillus chromosome. In some further embodiments, oneor more indigenous chromosomal regions have been deleted from acorresponding wild-type Bacillus host chromosome. In furtherembodiments, the Bacillus is an Alicyclobacillus sp. or Alicyclobacillusacidocaldarius.

Additional embodiments, include methods of modulating recombination attemperatures at or above about 25, 30, 35, 40, 45, 50, 55, 60, 65, 70,75, 80, 85, 90, and/or 95 degrees Celsius and/or at a pH at, below,and/or above 8, 7, 6, 5, 4, 3, 2, 1, and/or 0 via providing arecombinant, purified, and/or isolated nucleotide sequence comprising anucleotide sequence selected from the group consisting of a nucleotidesequences having at least 90% sequence identity to at least one of thesequences of SEQ ID Nos. 2, 19, 36, 53, 70, 87, 104, 121, 138, 155, 172,189, 206, 223, 240, 257, 274, 291, 308, 325, 359, 376, 410, 427, 444,512, 529, 546, 597, 614, 648, 665, 682, 699, 716, 733, 750, 767, 784,818, 835, 852, 886, 903, 920, 937, 954, 971, 988, 1005, 1039, 1056,1073, 1090, 1107, 1124, 1141, 1158, 1175, 1192, 1209, 1226, 1243, 1260,1277, 1294, 1311, 1328, 1345, 1362, 1379, 1396, 1413, 1430, 1447, 1464,1481, 1498, 1515, 1532, 1549, 1566, 1583, 1600, 1634, 1651, 1685, 1702,1719, 1787, 1804, 1821, 1872, 1889, 1923, 1940, 1957, 1974, 1991, 2008,2025, 2042, 2059, 2093, 2110, 2127, 2161, 2178, 2195, 2212, 2229, 2246,2263, 2280, 2314, 2331, 2348, 2365, 2382, 2399, 2416, 2433, 2450, 2467,2484, 2501, 2518, 2535, 2552, 2569, 2586, 2603, 2620, 2637, 2654, 2671,2688, 2705, 2722, 2739, 2773, 2790, 2824, 2841, 2858, 2926, 2943, 2960,3011, 3028, 3062, 3079, 3096, 3113, 3130, 3147, 3164, 3181, 3198, 3232,3249, 3266, and 3283; and/or a recombinant, purified, and/or isolatedpolypeptide selected from the group consisting of a polypeptide havingat least 90% sequence identity to at least one of the sequences of SEQID Nos. 1, 18, 35, 52, 69, 86, 103, 120, 137, 154, 171, 188, 205, 222,239, 256, 273, 290, 307, 324, 358, 375, 409, 426, 443, 511, 528, 545,596, 613, 647, 664, 681, 698, 715, 732, 749, 766, 783, 817, 834, 851,885, 902, 819, 936, 953, 970, 987, 1004, 1038, 1055, 1072, 1089, 1106,1123, 1140, 1157, 1174, 1191, 1208, 1225, 1242, 1259, 1276, 1293, 1310,1327, 1344, 1361, 1378, 1395, 1412, 1429, 1446, 1463, 1480, 1497, 1514,1531, 1548, 1565, 1582, 1599, 1633, 1650, 1684, 1701, 1718, 1786, 1803,1820, 1871, 1888, 1922, 1939, 1956, 1973, 1990, 2007, 2024, 2041, 2058,2092, 2109, 2126, 2160, 2177, 2094, 2211, 2228, 2245, 2262, 2279, 2313,2330, 2347, 2364, 2381, 2398, 2415, 2432, 2449, 2466, 2483, 2500, 2517,2534, 2551, 2568, 2585, 2602, 2619, 2636, 2653, 2670, 2687, 2704, 2721,2738, 2772, 2789, 2823, 2840, 2857, 2925, 2942, 2959, 3010, 3027, 3061,3078, 3095, 3112, 3129, 3146, 3163, 3180, 3197, 3231, 3248, 3265, and3282 to a nucleotide sequence in with which a recombination event isdesired.

In embodiments of the invention any one of the isolated and/or purifiedpolypeptides according to the invention may be enzymatically orfunctionally active at temperatures at or above about 25, 30, 35, 40,45, 50, 55, 60, 65, 70, 75, 80, 85, 90, and/or 95 degrees Celsius and/ormay be enzymatically or functionally active at a pH at, below, and/orabove 8, 7, 6, 5, 4, 3, 2, 1, and/or 0. In further embodiments of theinvention, glycosylation, pegylation, and/or other posttranslationalmodification may be required for the isolated and/or purifiedpolypeptides according to the invention to be enzymatically orfunctionally active at pH at or below 8, 7, 6, 5, 4, 3, 2, 1, and/or 0or at a temperatures at or above about 25, 30, 35, 40, 45, 50, 55, 60,65, 70, 75, 80, 85, 90, and/or 95 degrees Celsius.

The invention is described in additional detail in the followingillustrative examples. Although the examples may represent only selectedembodiments of the invention, it should be understood that the followingexamples are illustrative and not limiting.

EXAMPLES Example 1 Recombination Using Nucleotide and Amino AcidSequences from Alicyclobacillus acidocaldarius

Provided in SEQ ID Nos. 2, 19, 36, 53, 70, 87, 104, 121, 138, 155, 172,189, 206, 223, 240, 257, 274, 291, 308, 325, 359, 376, 410, 427, 444,512, 529, 546, 597, 614, 648, 665, 682, 699, 716, 733, 750, 767, 784,818, 835, 852, 886, 903, 920, 937, 954, 971, 988, 1005, 1039, 1056,1073, 1090, 1107, 1124, 1141, 1158, 1175, 1192, 1209, 1226, 1243, 1260,1277, 1294, 1311, 1328, 1345, 1362, 1379, 1396, 1413, 1430, 1447, 1464,1481, 1498, 1515, 1532, 1549, 1566, 1583, 1600, 1634, 1651, 1685, 1702,1719, 1787, 1804, 1821, 1872, 1889, 1923, 1940, 1957, 1974, 1991, 2008,2025, 2042, 2059, 2093, 2110, 2127, 2161, 2178, 2195, 2212, 2229, 2246,2263, 2280, 2314, 2331, 2348, 2365, 2382, 2399, 2416, 2433, 2450, 2467,2484, 2501, 2518, 2535, 2552, 2569, 2586, 2603, 2620, 2637, 2654, 2671,2688, 2705, 2722, 2739, 2773, 2790, 2824, 2841, 2858, 2926, 2943, 2960,3011, 3028, 3062, 3079, 3096, 3113, 3130, 3147, 3164, 3181, 3198, 3232,3249, 3266, and 3283 are a nucleotide sequences isolated fromAlicyclobacillus acidocaldarius and coding for the polypeptides of SEQID Nos. 1, 18, 35, 52, 69, 86, 103, 120, 137, 154, 171, 188, 205, 222,239, 256, 273, 290, 307, 324, 358, 375, 409, 426, 443, 511, 528, 545,596, 613, 647, 664, 681, 698, 715, 732, 749, 766, 783, 817, 834, 851,885, 902, 819, 936, 953, 970, 987, 1004, 1038, 1055, 1072, 1089, 1106,1123, 1140, 1157, 1174, 1191, 1208, 1225, 1242, 1259, 1276, 1293, 1310,1327, 1344, 1361, 1378, 1395, 1412, 1429, 1446, 1463, 1480, 1497, 1514,1531, 1548, 1565, 1582, 1599, 1633, 1650, 1684, 1701, 1718, 1786, 1803,1820, 1871, 1888, 1922, 1939, 1956, 1973, 1990, 2007, 2024, 2041, 2058,2092, 2109, 2126, 2160, 2177, 2094, 2211, 2228, 2245, 2262, 2279, 2313,2330, 2347, 2364, 2381, 2398, 2415, 2432, 2449, 2466, 2483, 2500, 2517,2534, 2551, 2568, 2585, 2602, 2619, 2636, 2653, 2670, 2687, 2704, 2721,2738, 2772, 2789, 2823, 2840, 2857, 2925, 2942, 2959, 3010, 3027, 3061,3078, 3095, 3112, 3129, 3146, 3163, 3180, 3197, 3231, 3248, 3265, and3282, and respectively. The nucleotide sequences of SEQ ID Nos. 2, 19,36, 53, 70, 87, 104, 121, 138, 155, 172, 189, 206, 223, 240, 257, 274,291, 308, 325, 359, 376, 410, 427, 444, 512, 529, 546, 597, 614, 648,665, 682, 699, 716, 733, 750, 767, 784, 818, 835, 852, 886, 903, 920,937, 954, 971, 988, 1005, 1039, 1056, 1073, 1090, 1107, 1124, 1141,1158, 1175, 1192, 1209, 1226, 1243, 1260, 1277, 1294, 1311, 1328, 1345,1362, 1379, 1396, 1413, 1430, 1447, 1464, 1481, 1498, 1515, 1532, 1549,1566, 1583, 1600, 1634, 1651, 1685, 1702, 1719, 1787, 1804, 1821, 1872,1889, 1923, 1940, 1957, 1974, 1991, 2008, 2025, 2042, 2059, 2093, 2110,2127, 2161, 2178, 2195, 2212, 2229, 2246, 2263, 2280, 2314, 2331, 2348,2365, 2382, 2399, 2416, 2433, 2450, 2467, 2484, 2501, 2518, 2535, 2552,2569, 2586, 2603, 2620, 2637, 2654, 2671, 2688, 2705, 2722, 2739, 2773,2790, 2824, 2841, 2858, 2926, 2943, 2960, 3011, 3028, 3062, 3079, 3096,3113, 3130, 3147, 3164, 3181, 3198, 3232, 3249, 3266, and 3283 areplaced into expression vectors using techniques standard in the art. Thevectors are then provided to cells such as bacteria cells or eukaryoticcells such as SD cells or CHO cells. In conjunction with the normalmachinery in present in the cells, the vectors comprising SEQ ID Nos. 2,19, 36, 53, 70, 87, 104, 121, 138, 155, 172, 189, 206, 223, 240, 257,274, 291, 308, 325, 359, 376, 410, 427, 444, 512, 529, 546, 597, 614,648, 665, 682, 699, 716, 733, 750, 767, 784, 818, 835, 852, 886, 903,920, 937, 954, 971, 988, 1005, 1039, 1056, 1073, 1090, 1107, 1124, 1141,1158, 1175, 1192, 1209, 1226, 1243, 1260, 1277, 1294, 1311, 1328, 1345,1362, 1379, 1396, 1413, 1430, 1447, 1464, 1481, 1498, 1515, 1532, 1549,1566, 1583, 1600, 1634, 1651, 1685, 1702, 1719, 1787, 1804, 1821, 1872,1889, 1923, 1940, 1957, 1974, 1991, 2008, 2025, 2042, 2059, 2093, 2110,2127, 2161, 2178, 2195, 2212, 2229, 2246, 2263, 2280, 2314, 2331, 2348,2365, 2382, 2399, 2416, 2433, 2450, 2467, 2484, 2501, 2518, 2535, 2552,2569, 2586, 2603, 2620, 2637, 2654, 2671, 2688, 2705, 2722, 2739, 2773,2790, 2824, 2841, 2858, 2926, 2943, 2960, 3011, 3028, 3062, 3079, 3096,3113, 3130, 3147, 3164, 3181, 3198, 3232, 3249, 3266, and 3283. Thepolypeptides of SEQ ID Nos. 1, 18, 35, 52, 69, 86, 103, 120, 137, 154,171, 188, 205, 222, 239, 256, 273, 290, 307, 324, 358, 375, 409, 426,443, 511, 528, 545, 596, 613, 647, 664, 681, 698, 715, 732, 749, 766,783, 817, 834, 851, 885, 902, 819, 936, 953, 970, 987, 1004, 1038, 1055,1072, 1089, 1106, 1123, 1140, 1157, 1174, 1191, 1208, 1225, 1242, 1259,1276, 1293, 1310, 1327, 1344, 1361, 1378, 1395, 1412, 1429, 1446, 1463,1480, 1497, 1514, 1531, 1548, 1565, 1582, 1599, 1633, 1650, 1684, 1701,1718, 1786, 1803, 1820, 1871, 1888, 1922, 1939, 1956, 1973, 1990, 2007,2024, 2041, 2058, 2092, 2109, 2126, 2160, 2177, 2094, 2211, 2228, 2245,2262, 2279, 2313, 2330, 2347, 2364, 2381, 2398, 2415, 2432, 2449, 2466,2483, 2500, 2517, 2534, 2551, 2568, 2585, 2602, 2619, 2636, 2653, 2670,2687, 2704, 2721, 2738, 2772, 2789, 2823, 2840, 2857, 2925, 2942, 2959,3010, 3027, 3061, 3078, 3095, 3112, 3129, 3146, 3163, 3180, 3197, 3231,3248, 3265, and 3282 are then isolated and/or purified. The isolatedand/or purified polypeptides of SEQ ID Nos. 1, 18, 35, 52, 69, 86, 103,120, 137, 154, 171, 188, 205, 222, 239, 256, 273, 290, 307, 324, 358,375, 409, 426, 443, 511, 528, 545, 596, 613, 647, 664, 681, 698, 715,732, 749, 766, 783, 817, 834, 851, 885, 902, 819, 936, 953, 970, 987,1004, 1038, 1055, 1072, 1089, 1106, 1123, 1140, 1157, 1174, 1191, 1208,1225, 1242, 1259, 1276, 1293, 1310, 1327, 1344, 1361, 1378, 1395, 1412,1429, 1446, 1463, 1480, 1497, 1514, 1531, 1548, 1565, 1582, 1599, 1633,1650, 1684, 1701, 1718, 1786, 1803, 1820, 1871, 1888, 1922, 1939, 1956,1973, 1990, 2007, 2024, 2041, 2058, 2092, 2109, 2126, 2160, 2177, 2094,2211, 2228, 2245, 2262, 2279, 2313, 2330, 2347, 2364, 2381, 2398, 2415,2432, 2449, 2466, 2483, 2500, 2517, 2534, 2551, 2568, 2585, 2602, 2619,2636, 2653, 2670, 2687, 2704, 2721, 2738, 2772, 2789, 2823, 2840, 2857,2925, 2942, 2959, 3010, 3027, 3061, 3078, 3095, 3112, 3129, 3146, 3163,3180, 3197, 3231, 3248, 3265, and 3282 are then each demonstrated tohave one or more of the activities provided in Table 1.

The isolated and/or purified polypeptides of SEQ ID Nos. 1, 18, 35, 52,69, 86, 103, 120, 137, 154, 171, 188, 205, 222, 239, 256, 273, 290, 307,324, 358, 375, 409, 426, 443, 511, 528, 545, 596, 613, 647, 664, 681,698, 715, 732, 749, 766, 783, 817, 834, 851, 885, 902, 819, 936, 953,970, 987, 1004, 1038, 1055, 1072, 1089, 1106, 1123, 1140, 1157, 1174,1191, 1208, 1225, 1242, 1259, 1276, 1293, 1310, 1327, 1344, 1361, 1378,1395, 1412, 1429, 1446, 1463, 1480, 1497, 1514, 1531, 1548, 1565, 1582,1599, 1633, 1650, 1684, 1701, 1718, 1786, 1803, 1820, 1871, 1888, 1922,1939, 1956, 1973, 1990, 2007, 2024, 2041, 2058, 2092, 2109, 2126, 2160,2177, 2094, 2211, 2228, 2245, 2262, 2279, 2313, 2330, 2347, 2364, 2381,2398, 2415, 2432, 2449, 2466, 2483, 2500, 2517, 2534, 2551, 2568, 2585,2602, 2619, 2636, 2653, 2670, 2687, 2704, 2721, 2738, 2772, 2789, 2823,2840, 2857, 2925, 2942, 2959, 3010, 3027, 3061, 3078, 3095, 3112, 3129,3146, 3163, 3180, 3197, 3231, 3248, 3265, and 3282 are demonstrated tohave activity as at least one of a ATP-dependent DNA helicase recG,ATP-dependent DNA ligase, ATP-dependent endopeptidase clp proteolyticsubunit, Chromosome partitioning protein, Crossover junctionendodeoxyribonuclease ruvC, Deoxyuridine 5′-triphosphatenucleotidohydrolase, DNA adenine methylase, DNA helicase, DNAintegration/recombination/inversion, DNA polymerase I, DNA polymeraseIII beta chain, DNA polymerase IV, DNA primase, DNA repair protein radC,DNA replication and repair protein recF, DNA replication protein dnaD,DNA topoisomerase I, DNA/RNA helicase (DEAD/DEAH box family),DNA-binding protein HU, Fe—S oxidoreductase, Glycerophosphoryl diesterphosphodiesterase, HNH endonuclease family protein, Holliday junctionDNA helicase ruvB, Integrase/recombinase (XerC/CodV family),Ligase/carboxyalse family protein, LtrC-like protein, Macrolide-effluxprotein, NAD-dependent DNA ligase, Nicotinate phosphoribosyltransferase,nodulin-26, Phage antirepressor protein, Phage protein, PhosphinothricinN-acetyltransferase, Phosphohydrolase, RecA protein, Recombinationprotein recR, Replicative DNA helicase, Ribonucleoside-diphosphatereductase alpha chain, Ribonucleoside-diphosphate reductase beta chain,Ribose-phosphate pyrophosphokinase, Serine/threonine proteinphosphatase, Single-strand DNA binding protein, Single-strandedDNA-binding protein, Site-specific recombinase, Site-specificresolvase/integrase, Thioredoxin, Thymidine kinase, Transcriptionalregulator/Lex A repressor, Transcriptional regulator, Cro/CI family,Transposase, TRSE protein, Two-component response regulator, Type IIrestriction-modification system methylation subunit.

All references, including publications, patents, and patentapplications, cited herein are hereby incorporated by reference to thesame extent as if each reference were individually and specificallyindicated to be incorporated by reference and were set forth in itsentirety herein.

While this invention has been described in certain embodiments, thepresent invention can be further modified within the spirit and scope ofthis disclosure. This application is therefore intended to cover anyvariations, uses, or adaptations of the invention using its generalprinciples. Further, this application is intended to cover suchdepartures from the present disclosure as come within known or customarypractice in the art to which this invention pertains and which fallwithin the limits of the appended claims and their legal equivalents.

BIBLIOGRAPHIC REFERENCES

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1. An isolated or purified nucleic acid sequence comprising a nucleicacid sequence encoding a polypeptide selected from the group consistingof a polypeptides having at least 90% sequence identity to SEQ ID Nos.1, 18, 35, 52, 69, 86, 103, 120, 137, 154, 171, 188, 205, 222, 239, 256,273, 290, 307, 324, 358, 375, 409, 426, 443, 511, 528, 545, 596, 613,647, 664, 681, 698, 715, 732, 749, 766, 783, 817, 834, 851, 885, 902,819, 936, 953, 970, 987, 1004, 1038, 1055, 1072, 1089, 1106, 1123, 1140,1157, 1174, 1191, 1208, 1225, 1242, 1259, 1276, 1293, 1310, 1327, 1344,1361, 1378, 1395, 1412, 1429, 1446, 1463, 1480, 1497, 1514, 1531, 1548,1565, 1582, 1599, 1633, 1650, 1684, 1701, 1718, 1786, 1803, 1820, 1871,1888, 1922, 1939, 1956, 1973, 1990, 2007, 2024, 2041, 2058, 2092, 2109,2126, 2160, 2177, 2094, 2211, 2228, 2245, 2262, 2279, 2313, 2330, 2347,2364, 2381, 2398, 2415, 2432, 2449, 2466, 2483, 2500, 2517, 2534, 2551,2568, 2585, 2602, 2619, 2636, 2653, 2670, 2687, 2704, 2721, 2738, 2772,2789, 2823, 2840, 2857, 2925, 2942, 2959, 3010, 3027, 3061, 3078, 3095,3112, 3129, 3146, 3163, 3180, 3197, 3231, 3248, 3265, and
 3282. 2. Theisolated or purified nucleic acid sequence of claim 1, wherein thepolypeptide has activity at or below about pH
 8. 3. The isolated orpurified nucleic acid sequence of claim 1, wherein the polypeptide hasactivity at a temperature at or above about 50 degrees Celsius.
 4. Theisolated or purified nucleic acid sequence of claim 1, wherein thenucleic acid sequence is present in a vector.
 5. An isolated or purifiedpolypeptide comprising a polypeptide selected from the group consistingof polypeptide having at least 90% sequence identity to SEQ ID Nos. 1,18, 35, 52, 69, 86, 103, 120, 137, 154, 171, 188, 205, 222, 239, 256,273, 290, 307, 324, 358, 375, 409, 426, 443, 511, 528, 545, 596, 613,647, 664, 681, 698, 715, 732, 749, 766, 783, 817, 834, 851, 885, 902,819, 936, 953, 970, 987, 1004, 1038, 1055, 1072, 1089, 1106, 1123, 1140,1157, 1174, 1191, 1208, 1225, 1242, 1259, 1276, 1293, 1310, 1327, 1344,1361, 1378, 1395, 1412, 1429, 1446, 1463, 1480, 1497, 1514, 1531, 1548,1565, 1582, 1599, 1633, 1650, 1684, 1701, 1718, 1786, 1803, 1820, 1871,1888, 1922, 1939, 1956, 1973, 1990, 2007, 2024, 2041, 2058, 2092, 2109,2126, 2160, 2177, 2094, 2211, 2228, 2245, 2262, 2279, 2313, 2330, 2347,2364, 2381, 2398, 2415, 2432, 2449, 2466, 2483, 2500, 2517, 2534, 2551,2568, 2585, 2602, 2619, 2636, 2653, 2670, 2687, 2704, 2721, 2738, 2772,2789, 2823, 2840, 2857, 2925, 2942, 2959, 3010, 3027, 3061, 3078, 3095,3112, 3129, 3146, 3163, 3180, 3197, 3231, 3248, 3265, and
 3282. 6. Theisolated or purified polypeptide of claim 5, wherein the polypeptide hasactivity at or below about pH
 8. 7. The isolated or purified polypeptideof claim 5, wherein the polypeptide has activity at a temperature at orabove about 50 degrees Celsius.
 8. The isolated or purified polypeptideof claim 5, wherein the polypeptide is glycosylated, pegylated, orotherwise post-translationally modified.
 9. The isolated or purifiedpolypeptide of claim 5, wherein the polypeptide has an activity selectedfrom the group consisting of ATP-dependent DNA helicase recG,ATP-dependent DNA ligase, ATP-dependent endopeptidase clp proteolyticsubunit, Chromosome partitioning protein, Crossover junctionendodeoxyribonuclease ruvC, Deoxyuridine 5′-triphosphatenucleotidohydrolase, DNA adenine methylase, DNA helicase, DNAintegration/recombination/inversion, DNA polymerase I, DNA polymeraseIII beta chain, DNA polymerase IV, DNA primase, DNA repair protein radC,DNA replication and repair protein recF, DNA replication protein dnaD,DNA topoisomerase I, DNA/RNA helicase (DEAD/DEAH box family),DNA-binding protein HU, Fe—S oxidoreductase, Glycerophosphoryl diesterphosphodiesterase, HNH endonuclease family protein, Holliday junctionDNA helicase ruvB, Integrase/recombinase (XerC/CodV family),Ligase/carboxyalse family protein, LtrC-like protein, Macrolide-effluxprotein, NAD-dependent DNA ligase, Nicotinate phosphoribosyltransferase,nodulin-26, Phage antirepressor protein, Phage protein, PhosphinothricinN-acetyltransferase, Phosphohydrolase, RecA protein, Recombinationprotein recR, Replicative DNA helicase, Ribonucleoside-diphosphatereductase alpha chain, Ribonucleoside-diphosphate reductase beta chain,Ribose-phosphate pyrophosphokinase, Serine/threonine proteinphosphatase, Single-strand DNA binding protein, Single-strandedDNA-binding protein, Site-specific recombinase, Site-specificresolvase/integrase, Thioredoxin, Thymidine kinase, Transcriptionalregulator/Lex A repressor, Transcriptional regulator, Cro/CI family,Transposase, TRSE protein, Two-component response regulator, Type IIrestriction-modification system methylation subunit activity.
 10. Amethod of modulating or altering recombination, the method comprising:providing a recombinant, purified, and/or isolated nucleotide sequencecomprising a nucleotide sequence selected from the group consisting of anucleotide sequences having at least 90% sequence identity to at leastone of the sequences of SEQ ID Nos. 2, 19, 36, 53, 70, 87, 104, 121,138, 155, 172, 189, 206, 223, 240, 257, 274, 291, 308, 325, 359, 376,410, 427, 444, 512, 529, 546, 597, 614, 648, 665, 682, 699, 716, 733,750, 767, 784, 818, 835, 852, 886, 903, 920, 937, 954, 971, 988, 1005,1039, 1056, 1073, 1090, 1107, 1124, 1141, 1158, 1175, 1192, 1209, 1226,1243, 1260, 1277, 1294, 1311, 1328, 1345, 1362, 1379, 1396, 1413, 1430,1447, 1464, 1481, 1498, 1515, 1532, 1549, 1566, 1583, 1600, 1634, 1651,1685, 1702, 1719, 1787, 1804, 1821, 1872, 1889, 1923, 1940, 1957, 1974,1991, 2008, 2025, 2042, 2059, 2093, 2110, 2127, 2161, 2178, 2195, 2212,2229, 2246, 2263, 2280, 2314, 2331, 2348, 2365, 2382, 2399, 2416, 2433,2450, 2467, 2484, 2501, 2518, 2535, 2552, 2569, 2586, 2603, 2620, 2637,2654, 2671, 2688, 2705, 2722, 2739, 2773, 2790, 2824, 2841, 2858, 2926,2943, 2960, 3011, 3028, 3062, 3079, 3096, 3113, 3130, 3147, 3164, 3181,3198, 3232, 3249, 3266, and 3283 to a sequence in which a geneticrecombination event is desired.
 11. The method according to claim 10,wherein the modulation or alteration of recombination occurs at or belowabout pH
 8. 12. The method according to claim 10, wherein the modulationor alteration of recombination occurs at a temperature at or above about50 degrees Celsius.
 13. The method according to claim 10, wherein therecombinant, purified, or isolated polypeptide is glycosylated,pegylated, or otherwise post-translationally modified.
 14. The methodaccording to claim 10, wherein the recombinant, purified, or isolatedpolypeptide has an activity selected from the group consisting ofATP-dependent DNA helicase recG, ATP-dependent DNA ligase, ATP-dependentendopeptidase clp proteolytic subunit, Chromosome partitioning protein,Crossover junction endodeoxyribonuclease ruvC, Deoxyuridine5′-triphosphate nucleotidohydrolase, DNA adenine methylase, DNAhelicase, DNA integration/recombination/inversion, DNA polymerase I, DNApolymerase III beta chain, DNA polymerase IV, DNA primase, DNA repairprotein radC, DNA replication and repair protein recF, DNA replicationprotein dnaD, DNA topoisomerase I, DNA/RNA helicase (DEAD/DEAH boxfamily), DNA-binding protein HU, Fe—S oxidoreductase, Glycerophosphoryldiester phosphodiesterase, HNH endonuclease family protein, Hollidayjunction DNA helicase ruvB, Integrase/recombinase (XerC/CodV family),Ligase/carboxyalse family protein, LtrC-like protein, Macrolide-effluxprotein, NAD-dependent DNA ligase, Nicotinate phosphoribosyltransferase,nodulin-26, Phage antirepressor protein, Phage protein, PhosphinothricinN-acetyltransferase, Phosphohydrolase, RecA protein, Recombinationprotein recR, Replicative DNA helicase, Ribonucleoside-diphosphatereductase alpha chain, Ribonucleoside-diphosphate reductase beta chain,Ribose-phosphate pyrophosphokinase, Serine/threonine proteinphosphatase, Single-strand DNA binding protein, Single-strandedDNA-binding protein, Site-specific recombinase, Site-specificresolvase/integrase, Thioredoxin, Thymidine kinase, Transcriptionalregulator/Lex A repressor, Transcriptional regulator, Cro/CI family,Transposase, TRSE protein, Two-component response regulator, Type IIrestriction-modification system methylation subunit activity.